Pia Huber - Academia.edu (original) (raw)

Papers by Pia Huber

Research paper thumbnail of The Binding of Distinct Segments of Actin to Multiple Sites in the C-Terminus of Caldesmon: Comparative Aspects of Actin Interaction with Troponin-I and Caldesmon

Biochemistry Usa, 1995

Thin-filament-based regulation of the contractile response is considered to involve the interacti... more Thin-filament-based regulation of the contractile response is considered to involve the interaction of actin with troponin-I in striated muscle and the interaction of actin with caldesmon in smooth muscle. The nature of the interaction with actin of these inhibitory proteins has been studied by proton magnetic resonance spectroscopy using segments of caldesmon and troponin-I which mimic their functional properties. Caldesmon is shown to interact with two distinct sites on the N-terminal residues 1-44 of actin subdomain 1 with corresponding contacts on caldesmon domain 3 and domain 4 at its C-terminus. We demonstrate that, whereas inhibition by the troponin-I fragment (residues 96-117) is effected by its interaction with the N-terminal region of actin, the separate inhibitory ability of different regions of the C-terminus of caldesmon (domains 4a and 4b) is mediated by interaction with noncontiguous segments on subdomain 1 of actin. Our studies of the spatial relationship of these actin contacts on caldesmon further suggest that one molecule of caldesmon may associate with two actin monomers. The demonstrated interactive nature of these caldesmon attachments to distinct regions of actin is relevant to the mechanism of calcium modulation of inhibition of actomyosin ATPase by caldesmon.

Research paper thumbnail of Conductive Education as a Method of Stroke Rehabilitation: A Single Blinded Randomised Controlled Feasibility Study

Stroke Research and Treatment, 2016

Background.Conductive Education for stroke survivors has shown promise but randomised evidence is... more Background.Conductive Education for stroke survivors has shown promise but randomised evidence is unavailable. This study assessed the feasibility of a definitive randomised controlled trial to evaluate efficacy.Methods.Adult stroke survivors were recruited through local community notices. Those completing the baseline assessment were randomised using an online program and group allocation was independent. Intervention group participants received 10 weekly 1.5-hour sessions of Conductive Education at the National Institute of Conductive Education in Birmingham, UK. The control group participants attended two group meetings. The study evaluated the feasibility of recruitment procedures, delivery of the intervention, retention of participants, and appropriateness of outcome measures and data collection methods. Independent assessments included the Barthel Index, the Stroke Impact Scale, the Timed Up and Go test, and the Hospital Anxiety and Depression Scale.Results.Eighty-two patients...

Research paper thumbnail of Disulfide formation within the regulatory light chain of skeletal muscle myosin

Biochemistry Usa, Dec 1, 1989

Thiol-disulfide exchange reactions between myosin and 5,5'-dithiobis(2-nitrobenzoic acid)... more Thiol-disulfide exchange reactions between myosin and 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) lead to the formation of 5-thio-2-nitrobenzoic acid (TNB)-mixed disulfides as well as to protein disulfide bonds. After incubation with DTNB, myosin was treated with an excess of N-ethylmaleimide (NEM) before electrophoretic analysis of the protein subunits in sodium dodecyl sulfate (SDS) without prior reduction by dithiothreitol (DTT). Without NEM treatment, thiol-disulfide rearrangement reactions occurred in the presence of SDS between the residual free thiols and DTNB. In the absence of divalent metal ions at 25 degrees C, DTNB was shown to induce an intrachain disulfide bond between Cys-127 and Cys-156 of the RLC. This intrachain cross-link restricts partially the unfolding of the RLC in SDS and can be followed as a faster migrating species, RLC'. Densitometric evaluation of the electrophoretic gel patterns indicated that the stoichiometric relation of the light chains (including RLC and RLC') remained unchanged. The two cysteine residues of the fast migrating RLC' were no more available for reaction with [14C]NEM, but upon reduction with DTT, the electrophoretic mobility of the RLC' reverted to that of unmodified RLC and of the RLC modified with two TNB groups. Ca2+ or Mg2+ was able to prevent this disulfide formation in the RLC of myosin by 50% at a free ion concentration of 1.1 X 10(-8) and 4.0 X 10(-7) M, respectively, at 25 degrees C and pH 7.6. Intrachain disulfide formation of RLC never occurred in myosin at 0 degree C.(ABSTRACT TRUNCATED AT 250 WORDS)

Research paper thumbnail of The Problem of Answered Prayer

Feminist Theology, 1996

I am a scientist by training and I offer my personal thoughts to a scientist and her article on p... more I am a scientist by training and I offer my personal thoughts to a scientist and her article on prayer.' Throughout my life I have had the chance to meet great personalities within religious circles. The inspiration from them has contributed substantially to my own religious ...

Research paper thumbnail of The Impact of National guidelines on Venom Immunotherapy Practice in the UK

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, Jan 17, 2016

Venom immunotherapy (VIT) is the only effective treatment for prevention of further anaphylactic ... more Venom immunotherapy (VIT) is the only effective treatment for prevention of further anaphylactic reactions to bee and wasp stings in allergic individuals. A previous national survey carried out in 2006/7 revealed significant heterogeneity in UK practice [1]. The British Society for Allergy and Clinical Immunology (BSACI) guidelines for the diagnosis and management of hymenoptera venom (HV) allergy were subsequently published in 2011 [2]. This repeat survey was carried out to assess whether publication of BSACI guidelines helped improve the diagnosis and management of HV allergy in the UK National Health Service (NHS). This article is protected by copyright. All rights reserved.

Research paper thumbnail of Practice of the Treatment of Milk Allergy in the UK: A National Audit

International archives of allergy and immunology, Jan 17, 2016

The management of cow's milk allergy (CMA) includes initial dairy exclusion with suitable die... more The management of cow's milk allergy (CMA) includes initial dairy exclusion with suitable dietary substitution, diagnostic challenges where indicated, and supervised re-introduction as the condition resolves. Information on clinician practice is limited. We audited the current practice of clinicians in the UK to evaluate adherence to international guidelines. An online questionnaire was sent to all clinicians whose practice included children identified on the national allergy society website. One hundred and sixty questionnaires were sent and 116 (73%) responded. Skin prick tests were more commonly used than serum-specific IgE as diagnostic tests. Respective proportions selecting amino acid formulas (AAF) or extensively hydrolysed formulas (EHF) in severe versus mild/moderate presentations were 78 and 40% versus 20 and 88%; soya formula was the first choice in 8.6 and 24%. The criteria for selection of AAF as the first choice were predominantly severe IgE-mediated and non-IgE-me...

Research paper thumbnail of Location of two contact sites between human caldesmon and Ca2+ calmodulin

Journal of Biological Chemistry

ABSTRACT

Research paper thumbnail of Struktur und Funktion der regulatorischen leichten Kette des Myosins im Skelettmuskel /

... Metadata Label, Value. Author(s): Huber, Pia Adelheid Julie. Publisher: Unknown. Citation:Hub... more ... Metadata Label, Value. Author(s): Huber, Pia Adelheid Julie. Publisher: Unknown. Citation:Huber, Pia Adelheid Julie. Struktur und Funktion der regulatorischen leichten Kette des Myosins im Skelettmuskel. (1990). doi:10.3929/ethz-a-000569261. ...

Research paper thumbnail of Prescription of Adrenaline Auto-Injectors - A National Survey of UK Allergy Specialists

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, Jan 19, 2015

Anaphylaxis is a life-threatening systemic reaction affecting all age groups prompting early admi... more Anaphylaxis is a life-threatening systemic reaction affecting all age groups prompting early administration of adrenaline (1, 2). Most cases of anaphylaxis occur in the community, thus underlining the importance for the provision of adrenaline auto-injector/s (AAIs), appropriate counselling and education for the patient (and carers where relevant) and school staff in children. Whilst foods are the commonest triggers in children, drugs, foods, idiopathic anaphylaxis (IA) and insect stings are common causes in adults (3). This article is protected by copyright. All rights reserved.

Research paper thumbnail of Multiple-sited interaction of caldesmon with Ca(2+)-calmodulin

The Biochemical journal, 1996

The binding of Ca(2+)- and Ba(2+)-calmodulin to caldesmon and its functional consequence was inve... more The binding of Ca(2+)- and Ba(2+)-calmodulin to caldesmon and its functional consequence was investigated with three different calmodulin mutants. Two calmodulin mutants have pairs of cysteine residues substituted and oxidized to a disulphide bond in either the N- or C-terminal lobe (C41/75 and C85/112). The third mutant has phenylalanine-92 replaced by alanine (F92A). Binding measurements in the presence of Ca2+ by separation on native gels and by carbodiimide-induced cross-linking showed a lower affinity for caldesmon in all the mutants. When Ca2+ was replaced by Ba2+ the affinity of calmodulin for caldesmon was further reduced. The ability of Ca(2+)-calmodulin to release caldesmon's inhibition of the actin-tropomyosin-activated myosin ATPase was virtually abolished by mutation of phenylalanine-92 to alanine or by replacing Ba2+ for Ca2+ in native calmodulin. Both cysteine mutants retained their functional ability, but the increased concentration needed for 50% release of cald...

Research paper thumbnail of BSACI guideline for the management of chronic urticaria and angioedema

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, 2015

This guidance for the management of patients with chronic urticaria and angioedema has been prepa... more This guidance for the management of patients with chronic urticaria and angioedema has been prepared by the Standards of Care Committee of the British Society for Allergy and Clinical Immunology (BSACI). The guideline is based on evidence as well as on expert opinion and is aimed at both adult physicians and paediatricians practising in allergy. The recommendations are evidence graded. During the development of these guidelines, all BSACI members were included in the consultation process using a Web-based system. Their comments and suggestions were carefully considered by the Standards of Care Committee. Where evidence was lacking, a consensus was reached by the experts on the committee. Included in this management guideline are clinical classification, aetiology, diagnosis, investigations, treatment guidance with special sections on children with urticaria and the use of antihistamines in women who are pregnant or breastfeeding. Finally, we have made recommendations for potential a...

Research paper thumbnail of Structural interactions between actin, tropomyosin, caldesmon and calcium binding protein and the regulation of smooth muscle thin filaments

Acta physiologica Scandinavica, 1998

The basic structure and functional properties of smooth muscle thin filaments were established ab... more The basic structure and functional properties of smooth muscle thin filaments were established about 10 years ago. Since then we and others have been working on the details of how tropomyosin, caldesmon and the Ca(2+)-binding protein regulate actin interaction with myosin. Our work has tended to emphasize the similarities between caldesmon and troponin function whilst others have been more concerned with the differences. The need to resolve the resulting differences has stimulated us to find new and more direct ways of investigating the mechanism of thin filament regulation. In recent years an apparent divergence has opened up between functional measurements, which indicate an allosteric-cooperative regulatory mechanism in which caldesmon and Ca(2+)-binding protein control actin-tropomyosin state in the same way as troponin, and structural measurements which show thin filament structures unlike striated muscle thin filaments. The challenge is to interpret function in terms of struct...

Research paper thumbnail of Location and functional characterization of myosin contact sites in smooth muscle caldesmon

The Biochemical journal, Jan 15, 1997

Caldesmon interaction with smooth muscle myosin and its ability to cross-link actin filaments to ... more Caldesmon interaction with smooth muscle myosin and its ability to cross-link actin filaments to myosin were investigated by the use of several bacterially expressed myosin-binding fragments of caldesmon. We have confirmed the presence of two functionally different myosin-binding sites located in domains 1 and 3/4a of caldesmon. The binding of the C-terminal site is highly sensitive to ionic strength and hardly participates in acto-myosin cross-linking, while the N-terminal binding site is relatively independent of ionic strength and apparently contains two separate myosin contact regions within residues 1-28 and 29-128 of chicken gizzard caldesmon. Both these N-terminal sub-sites are involved in the interaction with myosin and are predominantly responsible for the caldesmon-mediated high-affinity cross-linking of actin and myosin filaments, without affecting the affinity of direct acto-myosin interaction. Binding of caldesmon and its fragments to myosin or rod filaments revealed af...

Research paper thumbnail of Mapping of contact sites in the caldesmon-calmodulin complex

The Biochemical journal, Jan 15, 1997

The interaction of intact calmodulin and its four tryptic peptides with deletion mutants of calde... more The interaction of intact calmodulin and its four tryptic peptides with deletion mutants of caldesmon was analysed by native gel electrophoresis, fluorescence spectroscopy and zero-length cross-linking. Deletion mutants H2 (containing calmodulin-binding sites A and B) and H9 (containing sites B and B') interacted with intact calmodulin to form complexes whose stoichiometries varied from 2:1 to 1:1. The N-terminal peptides of calmodulin (TR1C, residues 1-77, and TR2E, residues 1-90) bound H2 with higher affinity than H9. At the same time H2 was less effective than H9 in binding to the C-terminal peptides of calmodulin TR2C (residues 78-148) and TR3E (residues 107-148). The N-terminal peptides of calmodulin (TR1C and TR2E) could be cross-linked to intact caldesmon and its deletion mutants H2 and H9. The similarity in the primary structures of sites A and B' of caldesmon and our measurements of the affinities of H2 and H9 to calmodulin and its peptides strongly indicate an orie...

Research paper thumbnail of Location of smooth-muscle myosin and tropomyosin binding sites in the C-terminal 288 residues of human caldesmon

The Biochemical journal, 1995

We have produced nine recombinant fragments, H1 to H9, from a human cDNA that codes for the C-ter... more We have produced nine recombinant fragments, H1 to H9, from a human cDNA that codes for the C-terminal 288 residues of caldesmon. The fragment H1, encompassing the 288 residues, is equivalent to domains 3 and 4 of caldesmon (amino acids 506-793 in human, 476-737 in the chicken gizzard sequence). It has been shown [Huber, Redwood, Avent, Tanner and Marston (1993) J. Muscle Res. Cell Motil. 14, 385-391] to bind to actin, Ca(2+)-calmodulin, tropomyosin and myosin. The fragments, H2 to H9, differ in length between 60 and 176 residues and cover the whole of domains 3 and 4 with many of the fragments overlapping. We have characterized the myosin and tropomyosin binding of these fragments. The binding of both tropomyosin and myosin is highly dependent on salt concentration, indicating the ionic nature of these interactions. The location of the myosin binding is an extended region encompassing the junction of domains 3/4 and domain 4a (residues 622-714, human; 566-657, chicken gizzard). Tro...

Research paper thumbnail of Smooth muscle caldesmon controls the strong binding interaction between actin-tropomyosin and myosin

The Journal of biological chemistry, Jan 23, 1994

We have demonstrated that caldesmon does not alter the affinity of weak binding actomyosin comple... more We have demonstrated that caldesmon does not alter the affinity of weak binding actomyosin complexes when it inhibits actin-tropomyosin activation at physiological ratios (1 per 14 actins), and we proposed that it acts upon the strong binding complexes in the same way that troponin-tropomyosin does. We therefore compared the effect of caldesmon, caldesmon fragments, and troponin upon the interaction of the strongly bound complexes S-1.ADP, S-1.adenylyl imidodiphosphate (AMP.PNP), and N-ethylmaleimide-treated myosin subfragment-1 (NEM-S-1) with actin-tropomyosin. In 0.17 M ionic strength buffer [14C]iodoacetamide-labeled S1.ADP bound to actin-smooth muscle tropomyosin with no evidence of cooperativity; Kd = 0.8 +/- 0.3 microM (n = 5). Inhibitory concentrations of sheep aorta caldesmon or rabbit skeletal muscle troponin made the binding highly cooperative. At low levels of saturation the apparent Kd was 10-40 microM with 10 microM caldesmon and 8-20 microM with 6 microM troponin; at &...

Research paper thumbnail of Caldesmon

Biochemistry of Smooth Muscle Contraction, 1996

Research paper thumbnail of Management of allergy to penicillins and other beta-lactams

Clinical & Experimental Allergy, 2015

The Standards of Care Committee of the British Society for Allergy and Clinical Immunology (BSACI... more The Standards of Care Committee of the British Society for Allergy and Clinical Immunology (BSACI) and an expert panel have prepared this guidance for the management of immediate and non-immediate allergic reactions to penicillins and other beta-lactams. The guideline is intended for UK specialists in both adult and paediatric allergy and for other clinicians practising allergy in secondary and tertiary care. The recommendations are evidence based, but where evidence is lacking, the panel reached consensus. During the development of the guideline, all BSACI members were consulted using a Web-based process and all comments carefully considered. Included in the guideline are epidemiology of allergic reactions to beta-lactams, molecular structure, formulations available in the UK and a description of known beta-lactam antigenic determinants. Sections on the value and limitations of clinical history, skin testing and laboratory investigations for both penicillins and cephalosporins are included. Cross-reactivity between penicillins and cephalosporins is discussed in detail. Recommendations on oral provocation and desensitization procedures have been made. Guidance for beta-lactam allergy in children is given in a separate section. An algorithm to help the clinician in the diagnosis of patients with a history of penicillin allergy has also been included.

Research paper thumbnail of Identification of functioning regulatory sites and a new myosin binding site in the C-terminal 288 amino acids of caldesmon expressed from a human clone

Journal of Muscle Research and Cell Motility, 1993

A partial clone of caldesmon, coding for the C-terminal 288 amino acids, was isolated from a huma... more A partial clone of caldesmon, coding for the C-terminal 288 amino acids, was isolated from a human fetal liver cDNA library and sequenced. Expression of the clone in Escherichia coli produced a peptide called H1 (M r 32 549), which inhibited tropomyosin-enhanced actomyosin Mg2+-ATPase activity by 90% with half maximal inhibition at 0.03-0.04 mol H1 per mol actin. The inhibition could be reversed by Ca2+-calmodulin. H1 bound actin, Ca2+-calmodulin and tropomyosin and smooth muscle myosin with high affinities. This latter finding shows the presence of a second myosin-binding site in caldesmon. This was confirmed in thrombic digests of native sheep aorta and chicken gizzard caldesmon.

Research paper thumbnail of Abstracts of the XVIII European Conference on Muscle and Motility

Journal of Muscle Research and Cell Motility, 1990

Four major inorganic cations--Na+, K--, Mg 2+ and Ca 2+ contribute mainly to the regulation of ac... more Four major inorganic cations--Na+, K--, Mg 2+ and Ca 2+ contribute mainly to the regulation of activity of muscle cells. The aim of the present comparative study was fo reveal the principal factors which determine the great variety of the cationic contents in different ...

Research paper thumbnail of The Binding of Distinct Segments of Actin to Multiple Sites in the C-Terminus of Caldesmon: Comparative Aspects of Actin Interaction with Troponin-I and Caldesmon

Biochemistry Usa, 1995

Thin-filament-based regulation of the contractile response is considered to involve the interacti... more Thin-filament-based regulation of the contractile response is considered to involve the interaction of actin with troponin-I in striated muscle and the interaction of actin with caldesmon in smooth muscle. The nature of the interaction with actin of these inhibitory proteins has been studied by proton magnetic resonance spectroscopy using segments of caldesmon and troponin-I which mimic their functional properties. Caldesmon is shown to interact with two distinct sites on the N-terminal residues 1-44 of actin subdomain 1 with corresponding contacts on caldesmon domain 3 and domain 4 at its C-terminus. We demonstrate that, whereas inhibition by the troponin-I fragment (residues 96-117) is effected by its interaction with the N-terminal region of actin, the separate inhibitory ability of different regions of the C-terminus of caldesmon (domains 4a and 4b) is mediated by interaction with noncontiguous segments on subdomain 1 of actin. Our studies of the spatial relationship of these actin contacts on caldesmon further suggest that one molecule of caldesmon may associate with two actin monomers. The demonstrated interactive nature of these caldesmon attachments to distinct regions of actin is relevant to the mechanism of calcium modulation of inhibition of actomyosin ATPase by caldesmon.

Research paper thumbnail of Conductive Education as a Method of Stroke Rehabilitation: A Single Blinded Randomised Controlled Feasibility Study

Stroke Research and Treatment, 2016

Background.Conductive Education for stroke survivors has shown promise but randomised evidence is... more Background.Conductive Education for stroke survivors has shown promise but randomised evidence is unavailable. This study assessed the feasibility of a definitive randomised controlled trial to evaluate efficacy.Methods.Adult stroke survivors were recruited through local community notices. Those completing the baseline assessment were randomised using an online program and group allocation was independent. Intervention group participants received 10 weekly 1.5-hour sessions of Conductive Education at the National Institute of Conductive Education in Birmingham, UK. The control group participants attended two group meetings. The study evaluated the feasibility of recruitment procedures, delivery of the intervention, retention of participants, and appropriateness of outcome measures and data collection methods. Independent assessments included the Barthel Index, the Stroke Impact Scale, the Timed Up and Go test, and the Hospital Anxiety and Depression Scale.Results.Eighty-two patients...

Research paper thumbnail of Disulfide formation within the regulatory light chain of skeletal muscle myosin

Biochemistry Usa, Dec 1, 1989

Thiol-disulfide exchange reactions between myosin and 5,5'-dithiobis(2-nitrobenzoic acid)... more Thiol-disulfide exchange reactions between myosin and 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) lead to the formation of 5-thio-2-nitrobenzoic acid (TNB)-mixed disulfides as well as to protein disulfide bonds. After incubation with DTNB, myosin was treated with an excess of N-ethylmaleimide (NEM) before electrophoretic analysis of the protein subunits in sodium dodecyl sulfate (SDS) without prior reduction by dithiothreitol (DTT). Without NEM treatment, thiol-disulfide rearrangement reactions occurred in the presence of SDS between the residual free thiols and DTNB. In the absence of divalent metal ions at 25 degrees C, DTNB was shown to induce an intrachain disulfide bond between Cys-127 and Cys-156 of the RLC. This intrachain cross-link restricts partially the unfolding of the RLC in SDS and can be followed as a faster migrating species, RLC'. Densitometric evaluation of the electrophoretic gel patterns indicated that the stoichiometric relation of the light chains (including RLC and RLC') remained unchanged. The two cysteine residues of the fast migrating RLC' were no more available for reaction with [14C]NEM, but upon reduction with DTT, the electrophoretic mobility of the RLC' reverted to that of unmodified RLC and of the RLC modified with two TNB groups. Ca2+ or Mg2+ was able to prevent this disulfide formation in the RLC of myosin by 50% at a free ion concentration of 1.1 X 10(-8) and 4.0 X 10(-7) M, respectively, at 25 degrees C and pH 7.6. Intrachain disulfide formation of RLC never occurred in myosin at 0 degree C.(ABSTRACT TRUNCATED AT 250 WORDS)

Research paper thumbnail of The Problem of Answered Prayer

Feminist Theology, 1996

I am a scientist by training and I offer my personal thoughts to a scientist and her article on p... more I am a scientist by training and I offer my personal thoughts to a scientist and her article on prayer.' Throughout my life I have had the chance to meet great personalities within religious circles. The inspiration from them has contributed substantially to my own religious ...

Research paper thumbnail of The Impact of National guidelines on Venom Immunotherapy Practice in the UK

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, Jan 17, 2016

Venom immunotherapy (VIT) is the only effective treatment for prevention of further anaphylactic ... more Venom immunotherapy (VIT) is the only effective treatment for prevention of further anaphylactic reactions to bee and wasp stings in allergic individuals. A previous national survey carried out in 2006/7 revealed significant heterogeneity in UK practice [1]. The British Society for Allergy and Clinical Immunology (BSACI) guidelines for the diagnosis and management of hymenoptera venom (HV) allergy were subsequently published in 2011 [2]. This repeat survey was carried out to assess whether publication of BSACI guidelines helped improve the diagnosis and management of HV allergy in the UK National Health Service (NHS). This article is protected by copyright. All rights reserved.

Research paper thumbnail of Practice of the Treatment of Milk Allergy in the UK: A National Audit

International archives of allergy and immunology, Jan 17, 2016

The management of cow's milk allergy (CMA) includes initial dairy exclusion with suitable die... more The management of cow's milk allergy (CMA) includes initial dairy exclusion with suitable dietary substitution, diagnostic challenges where indicated, and supervised re-introduction as the condition resolves. Information on clinician practice is limited. We audited the current practice of clinicians in the UK to evaluate adherence to international guidelines. An online questionnaire was sent to all clinicians whose practice included children identified on the national allergy society website. One hundred and sixty questionnaires were sent and 116 (73%) responded. Skin prick tests were more commonly used than serum-specific IgE as diagnostic tests. Respective proportions selecting amino acid formulas (AAF) or extensively hydrolysed formulas (EHF) in severe versus mild/moderate presentations were 78 and 40% versus 20 and 88%; soya formula was the first choice in 8.6 and 24%. The criteria for selection of AAF as the first choice were predominantly severe IgE-mediated and non-IgE-me...

Research paper thumbnail of Location of two contact sites between human caldesmon and Ca2+ calmodulin

Journal of Biological Chemistry

ABSTRACT

Research paper thumbnail of Struktur und Funktion der regulatorischen leichten Kette des Myosins im Skelettmuskel /

... Metadata Label, Value. Author(s): Huber, Pia Adelheid Julie. Publisher: Unknown. Citation:Hub... more ... Metadata Label, Value. Author(s): Huber, Pia Adelheid Julie. Publisher: Unknown. Citation:Huber, Pia Adelheid Julie. Struktur und Funktion der regulatorischen leichten Kette des Myosins im Skelettmuskel. (1990). doi:10.3929/ethz-a-000569261. ...

Research paper thumbnail of Prescription of Adrenaline Auto-Injectors - A National Survey of UK Allergy Specialists

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, Jan 19, 2015

Anaphylaxis is a life-threatening systemic reaction affecting all age groups prompting early admi... more Anaphylaxis is a life-threatening systemic reaction affecting all age groups prompting early administration of adrenaline (1, 2). Most cases of anaphylaxis occur in the community, thus underlining the importance for the provision of adrenaline auto-injector/s (AAIs), appropriate counselling and education for the patient (and carers where relevant) and school staff in children. Whilst foods are the commonest triggers in children, drugs, foods, idiopathic anaphylaxis (IA) and insect stings are common causes in adults (3). This article is protected by copyright. All rights reserved.

Research paper thumbnail of Multiple-sited interaction of caldesmon with Ca(2+)-calmodulin

The Biochemical journal, 1996

The binding of Ca(2+)- and Ba(2+)-calmodulin to caldesmon and its functional consequence was inve... more The binding of Ca(2+)- and Ba(2+)-calmodulin to caldesmon and its functional consequence was investigated with three different calmodulin mutants. Two calmodulin mutants have pairs of cysteine residues substituted and oxidized to a disulphide bond in either the N- or C-terminal lobe (C41/75 and C85/112). The third mutant has phenylalanine-92 replaced by alanine (F92A). Binding measurements in the presence of Ca2+ by separation on native gels and by carbodiimide-induced cross-linking showed a lower affinity for caldesmon in all the mutants. When Ca2+ was replaced by Ba2+ the affinity of calmodulin for caldesmon was further reduced. The ability of Ca(2+)-calmodulin to release caldesmon's inhibition of the actin-tropomyosin-activated myosin ATPase was virtually abolished by mutation of phenylalanine-92 to alanine or by replacing Ba2+ for Ca2+ in native calmodulin. Both cysteine mutants retained their functional ability, but the increased concentration needed for 50% release of cald...

Research paper thumbnail of BSACI guideline for the management of chronic urticaria and angioedema

Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology, 2015

This guidance for the management of patients with chronic urticaria and angioedema has been prepa... more This guidance for the management of patients with chronic urticaria and angioedema has been prepared by the Standards of Care Committee of the British Society for Allergy and Clinical Immunology (BSACI). The guideline is based on evidence as well as on expert opinion and is aimed at both adult physicians and paediatricians practising in allergy. The recommendations are evidence graded. During the development of these guidelines, all BSACI members were included in the consultation process using a Web-based system. Their comments and suggestions were carefully considered by the Standards of Care Committee. Where evidence was lacking, a consensus was reached by the experts on the committee. Included in this management guideline are clinical classification, aetiology, diagnosis, investigations, treatment guidance with special sections on children with urticaria and the use of antihistamines in women who are pregnant or breastfeeding. Finally, we have made recommendations for potential a...

Research paper thumbnail of Structural interactions between actin, tropomyosin, caldesmon and calcium binding protein and the regulation of smooth muscle thin filaments

Acta physiologica Scandinavica, 1998

The basic structure and functional properties of smooth muscle thin filaments were established ab... more The basic structure and functional properties of smooth muscle thin filaments were established about 10 years ago. Since then we and others have been working on the details of how tropomyosin, caldesmon and the Ca(2+)-binding protein regulate actin interaction with myosin. Our work has tended to emphasize the similarities between caldesmon and troponin function whilst others have been more concerned with the differences. The need to resolve the resulting differences has stimulated us to find new and more direct ways of investigating the mechanism of thin filament regulation. In recent years an apparent divergence has opened up between functional measurements, which indicate an allosteric-cooperative regulatory mechanism in which caldesmon and Ca(2+)-binding protein control actin-tropomyosin state in the same way as troponin, and structural measurements which show thin filament structures unlike striated muscle thin filaments. The challenge is to interpret function in terms of struct...

Research paper thumbnail of Location and functional characterization of myosin contact sites in smooth muscle caldesmon

The Biochemical journal, Jan 15, 1997

Caldesmon interaction with smooth muscle myosin and its ability to cross-link actin filaments to ... more Caldesmon interaction with smooth muscle myosin and its ability to cross-link actin filaments to myosin were investigated by the use of several bacterially expressed myosin-binding fragments of caldesmon. We have confirmed the presence of two functionally different myosin-binding sites located in domains 1 and 3/4a of caldesmon. The binding of the C-terminal site is highly sensitive to ionic strength and hardly participates in acto-myosin cross-linking, while the N-terminal binding site is relatively independent of ionic strength and apparently contains two separate myosin contact regions within residues 1-28 and 29-128 of chicken gizzard caldesmon. Both these N-terminal sub-sites are involved in the interaction with myosin and are predominantly responsible for the caldesmon-mediated high-affinity cross-linking of actin and myosin filaments, without affecting the affinity of direct acto-myosin interaction. Binding of caldesmon and its fragments to myosin or rod filaments revealed af...

Research paper thumbnail of Mapping of contact sites in the caldesmon-calmodulin complex

The Biochemical journal, Jan 15, 1997

The interaction of intact calmodulin and its four tryptic peptides with deletion mutants of calde... more The interaction of intact calmodulin and its four tryptic peptides with deletion mutants of caldesmon was analysed by native gel electrophoresis, fluorescence spectroscopy and zero-length cross-linking. Deletion mutants H2 (containing calmodulin-binding sites A and B) and H9 (containing sites B and B') interacted with intact calmodulin to form complexes whose stoichiometries varied from 2:1 to 1:1. The N-terminal peptides of calmodulin (TR1C, residues 1-77, and TR2E, residues 1-90) bound H2 with higher affinity than H9. At the same time H2 was less effective than H9 in binding to the C-terminal peptides of calmodulin TR2C (residues 78-148) and TR3E (residues 107-148). The N-terminal peptides of calmodulin (TR1C and TR2E) could be cross-linked to intact caldesmon and its deletion mutants H2 and H9. The similarity in the primary structures of sites A and B' of caldesmon and our measurements of the affinities of H2 and H9 to calmodulin and its peptides strongly indicate an orie...

Research paper thumbnail of Location of smooth-muscle myosin and tropomyosin binding sites in the C-terminal 288 residues of human caldesmon

The Biochemical journal, 1995

We have produced nine recombinant fragments, H1 to H9, from a human cDNA that codes for the C-ter... more We have produced nine recombinant fragments, H1 to H9, from a human cDNA that codes for the C-terminal 288 residues of caldesmon. The fragment H1, encompassing the 288 residues, is equivalent to domains 3 and 4 of caldesmon (amino acids 506-793 in human, 476-737 in the chicken gizzard sequence). It has been shown [Huber, Redwood, Avent, Tanner and Marston (1993) J. Muscle Res. Cell Motil. 14, 385-391] to bind to actin, Ca(2+)-calmodulin, tropomyosin and myosin. The fragments, H2 to H9, differ in length between 60 and 176 residues and cover the whole of domains 3 and 4 with many of the fragments overlapping. We have characterized the myosin and tropomyosin binding of these fragments. The binding of both tropomyosin and myosin is highly dependent on salt concentration, indicating the ionic nature of these interactions. The location of the myosin binding is an extended region encompassing the junction of domains 3/4 and domain 4a (residues 622-714, human; 566-657, chicken gizzard). Tro...

Research paper thumbnail of Smooth muscle caldesmon controls the strong binding interaction between actin-tropomyosin and myosin

The Journal of biological chemistry, Jan 23, 1994

We have demonstrated that caldesmon does not alter the affinity of weak binding actomyosin comple... more We have demonstrated that caldesmon does not alter the affinity of weak binding actomyosin complexes when it inhibits actin-tropomyosin activation at physiological ratios (1 per 14 actins), and we proposed that it acts upon the strong binding complexes in the same way that troponin-tropomyosin does. We therefore compared the effect of caldesmon, caldesmon fragments, and troponin upon the interaction of the strongly bound complexes S-1.ADP, S-1.adenylyl imidodiphosphate (AMP.PNP), and N-ethylmaleimide-treated myosin subfragment-1 (NEM-S-1) with actin-tropomyosin. In 0.17 M ionic strength buffer [14C]iodoacetamide-labeled S1.ADP bound to actin-smooth muscle tropomyosin with no evidence of cooperativity; Kd = 0.8 +/- 0.3 microM (n = 5). Inhibitory concentrations of sheep aorta caldesmon or rabbit skeletal muscle troponin made the binding highly cooperative. At low levels of saturation the apparent Kd was 10-40 microM with 10 microM caldesmon and 8-20 microM with 6 microM troponin; at &...

Research paper thumbnail of Caldesmon

Biochemistry of Smooth Muscle Contraction, 1996

Research paper thumbnail of Management of allergy to penicillins and other beta-lactams

Clinical & Experimental Allergy, 2015

The Standards of Care Committee of the British Society for Allergy and Clinical Immunology (BSACI... more The Standards of Care Committee of the British Society for Allergy and Clinical Immunology (BSACI) and an expert panel have prepared this guidance for the management of immediate and non-immediate allergic reactions to penicillins and other beta-lactams. The guideline is intended for UK specialists in both adult and paediatric allergy and for other clinicians practising allergy in secondary and tertiary care. The recommendations are evidence based, but where evidence is lacking, the panel reached consensus. During the development of the guideline, all BSACI members were consulted using a Web-based process and all comments carefully considered. Included in the guideline are epidemiology of allergic reactions to beta-lactams, molecular structure, formulations available in the UK and a description of known beta-lactam antigenic determinants. Sections on the value and limitations of clinical history, skin testing and laboratory investigations for both penicillins and cephalosporins are included. Cross-reactivity between penicillins and cephalosporins is discussed in detail. Recommendations on oral provocation and desensitization procedures have been made. Guidance for beta-lactam allergy in children is given in a separate section. An algorithm to help the clinician in the diagnosis of patients with a history of penicillin allergy has also been included.

Research paper thumbnail of Identification of functioning regulatory sites and a new myosin binding site in the C-terminal 288 amino acids of caldesmon expressed from a human clone

Journal of Muscle Research and Cell Motility, 1993

A partial clone of caldesmon, coding for the C-terminal 288 amino acids, was isolated from a huma... more A partial clone of caldesmon, coding for the C-terminal 288 amino acids, was isolated from a human fetal liver cDNA library and sequenced. Expression of the clone in Escherichia coli produced a peptide called H1 (M r 32 549), which inhibited tropomyosin-enhanced actomyosin Mg2+-ATPase activity by 90% with half maximal inhibition at 0.03-0.04 mol H1 per mol actin. The inhibition could be reversed by Ca2+-calmodulin. H1 bound actin, Ca2+-calmodulin and tropomyosin and smooth muscle myosin with high affinities. This latter finding shows the presence of a second myosin-binding site in caldesmon. This was confirmed in thrombic digests of native sheep aorta and chicken gizzard caldesmon.

Research paper thumbnail of Abstracts of the XVIII European Conference on Muscle and Motility

Journal of Muscle Research and Cell Motility, 1990

Four major inorganic cations--Na+, K--, Mg 2+ and Ca 2+ contribute mainly to the regulation of ac... more Four major inorganic cations--Na+, K--, Mg 2+ and Ca 2+ contribute mainly to the regulation of activity of muscle cells. The aim of the present comparative study was fo reveal the principal factors which determine the great variety of the cationic contents in different ...