kalman buki | University of Turku (original) (raw)
Papers by kalman buki
Phytochemistry, 1972
A pyridoxal phosphate-dependent tryptophan decarboxylase has been purified 20-fold from seedlings... more A pyridoxal phosphate-dependent tryptophan decarboxylase has been purified 20-fold from seedlings of Phaluris tuberosa. The enzyme activity of the seedlings reached a maximum after 4 days. The enzyme activity is maximal at pH 7.6 and could be demonstrated either by the production of r4C-tryptamine from methylene 14C-L-tryptophan or of 14C02 from carboxyl-r4C-L-tryptophan. o-tryptophan was not a substrate but 5-hydroxy-14C-tryptophan was. The decarboxylation was inhibited at concentrations of 0.1 mM by a number of indole derivatives including N,N-dimethyltryptamine, one of the two major alkaloids produced by the plant. The second alkaloid, 5-methoxy-N,N-dimethyltryptamine did not inhibit at this concentration. A possible role of this enzyme in alkaloid biosynthesis is discussed.
PubMed, 1987
The proliferation stimulating or inhibitory effect of aortic endothelial cells on aortic smooth m... more The proliferation stimulating or inhibitory effect of aortic endothelial cells on aortic smooth muscle cells and vice versa was studied in cell culture by using a newly developed co-cultivation technique. The effect was registered as the incorporation of labelled thymidine detected by liquid scintillation counting. The experiments were performed in intra and interspecific combinations, using bovine, pig or rat aortic cell cultures. The endothelial cells stimulated smooth muscle cell proliferation in a species specific way. In interspecies combinations either inhibition or no effect was observed. The smooth muscle cells usually had an inhibitory effect on the proliferation of endothelial cells except for the pig smooth muscle cells which had a stimulatory effect in both inter and intraspecies combinations. Co-cultivation of smooth muscle or endothelial cells with cells of the same type resulted mostly in inhibition or no effect in both intra and interspecies combinations. This phenomenon was particularly conspicuous with endothelial cells. The different stimulatory and inhibitory effects produced by these cells may play an important role in the regulation of neovascularization and in reendothelization of denuded intimal areas.
PubMed, 1986
[U-14C]-glucose was incorporated non-enzymatically into the protein moiety of human low density l... more [U-14C]-glucose was incorporated non-enzymatically into the protein moiety of human low density lipoprotein. The incorporation was time and glucose concentration dependent. Investigated mini pig aortic endothelial cells showed that the glycosylated low density lipoprotein was bound, internalized and degraded significantly less than that of the native one measured with [125-I]-low density lipoprotein.
Differentiation, Dec 4, 2016
Fam3c, a cytokine-like protein, is a member of the Fam3 family (family with sequence similarity 3... more Fam3c, a cytokine-like protein, is a member of the Fam3 family (family with sequence similarity 3) and has been implicated to play a crucial role in Epithelial-to-mesenchymal transition (EMT) and subsequent metastasis during cancer progression. A few independent genome-wide association studies on different population cohorts predicted the gene locus of Fam3c to be associated with bone mineral density and fractures. In this study, we examined the role of Fam3c during osteoblast differentiation. Fam3c was found to be expressed during osteogenic differentiation of both primary bone marrow stromal cells and MC3T3-E1 pre-osteoblasts. In differentiating osteoblasts, knockdown of Fam3c increased alkaline phosphatase expression and activity whereas overexpression of Fam3c reduced it. Furthermore, overexpression of Fam3c caused reduction of Runx2 expression at both mRNA and protein levels. Fam3c was localized in the cytoplasm and it was not secreted outside the cell during osteoblast differentiation and therefore, may function intracellularly. Furthermore, Fam3c and TGF-β1 were found to regulate each other reciprocally. Our findings therefore suggest a functional role of Fam3c in the regulation of osteoblast differentiation. 2. Materials and methods 2.1. Cell culture MC3T3-E1 cells were cultured in α-MEM without ascorbic acid,
ADP-ribosyl transferase is a specific DNA-associating nuclear protein, whose enzymatic activity d... more ADP-ribosyl transferase is a specific DNA-associating nuclear protein, whose enzymatic activity depends on DNA-binding, probably to certain DNA sequences (1, 2). The study of molecular and biochemical mechanisms involving ADP-ribosyl transferase depends on experimentally efficient isolation of the enzyme. We present here a method that yields 8–9 mg of 95% homogeneous enzyme per kg starting material (calf thymus) within 3 days. In order to determine the polypeptide structure of ADP- ribosyl transferase, (with special reference to catalytically active and binding domains), we prepared stable polypeptides from ADP-ribosyl transferase by digestion with plasmin. We also report here on the partial sequencing of a polypeptide (36 kDa) that participates in the DNA-binding function of ADP- ribosyl transferase.
International Journal of Oncology, Dec 1, 1997
The GTPase activity of purified dimeric tubulin (alpha+beta) at 5 mu M was insensitive to methyl-... more The GTPase activity of purified dimeric tubulin (alpha+beta) at 5 mu M was insensitive to methyl-3,5-diiodo-4-(4'-methoxyphenoxy) benzoate (DIME), in contrast to nocodazole which activated GTPase. Cellular motility of MDA-MB-231 (human mammary cancer) cells migrating through 12-mu m pores was inhibited by DIME similar to nocodazole in a drug concentration-and DIME structure-dependent manner. An increase of cytoplasmic ATPase activity of DIME-treated cells without a decrease in ATP contents of intact cells suggests that DIME may also influence additional as yet unidentified ATP-dependent system(s) probably also involved in cell motility. These results show that DIME not only arrests cells in M phase but also inhibits cell motility in interphase. However the cellular mode of action of DIME is different from the action of other toxic tubulin-targeted drugs, despite the fact that DIME in a concentration-dependent manner disrupts microtubule structures in intact cells.
PubMed, 1990
To clarify whether some viruses could influence the different functions and membrane permeability... more To clarify whether some viruses could influence the different functions and membrane permeability of the aortic cells, we have examined in a model experiment the in vitro effect of the measles virus on the aortic endothelial and smooth muscle cells. The aortic cells infected with the virus failed to reveal gross cytopathic effect. Occasionally, however, syncytium formation and nuclear inclusions were observed. In infected endothelial cells lysosome containing viral nucleocapsids were seen. The early phase of measles virus replication inhibited the proliferation of endothelial cells of all species tested, while uniformly stimulated the replication of the smooth muscle cells relative to the control. In bovine aortic endothelial and smooth muscle cells the protein synthesis had been suppressed by the 4th to 6th hours postinfection. The results indicate that measles virus infection may be among the risk factors of atherosclerosis. It may damage endothelial cells by altering the cell membrane permeability and could induce proliferation of aortic smooth muscle cells.
Thrombosis and Haemostasis, 1983
SummaryHeparin forms a complex with human low density lipoprotein (LDL) in the presence of Ca2+. ... more SummaryHeparin forms a complex with human low density lipoprotein (LDL) in the presence of Ca2+. The complex is dissociable by 0.5 M NaCI. Thrombin and plasmin causes the dissociation of the LDL-heparin complex, whereas factor Xa does not. Heparin, complexed with LDL, retains its enhancing effect on the rate of thrombin and plasmin inactivation by antithrombin III. LDL isolated from the plasma of persons with different pathological conditions did not alter the rate of thrombin inactivation by antithrombin III either in the absence or in the presence of heparin. Heparin seems to maintain its biological functions when it is in a complex with LDL.
Molecular and Cellular Endocrinology, 2017
Fibroblast growth factors (FGF) and their receptors (FGFRs) regulate many developmental processes... more Fibroblast growth factors (FGF) and their receptors (FGFRs) regulate many developmental processes including differentiation of mesenchymal stromal cells (MSC). We developed two MSC lines capable of differentiating to osteoblasts and adipocytes and studied the role of FGFRs in this process. We identified FGFR2 and fibroblast growth factor receptor like-1 (FGFRL1) as possible actors in MSC differentiation with gene microarray and qRT-PCR. FGFR2 and FGFRL1 mRNA expression strongly increased during MSC differentiation to osteoblasts. FGF2 treatment, resulting in downregulation of FGFR2, or silencing FGFR2 expression with siRNAs inhibited osteoblast differentiation. During adipocyte differentiation expression of FGFR1 and FGFRL1 increased and was down-regulated by FGF2. FGFR1 knockdown inhibited adipocyte differentiation. Silencing FGFR2 and FGFR1 in MSCs was associated with decreased FGFRL1 expression in osteoblasts and adipocytes, respectively. Our results suggest that FGFR1 and FGFR2 regulate FGFRL1 expression. FGFRL1 may mediate or modulate FGFR regulation of MSC differentiation together with FGFR2 in osteoblastic and FGFR1 in adipocytic lineage.
Biochemical Journal, 1984
Interaction of human plasmin with a monolayer culture of mini-pig aortic endothelial cells was st... more Interaction of human plasmin with a monolayer culture of mini-pig aortic endothelial cells was studied by using the 125I-labelled enzyme. The binding of plasmin was time- and concentration-dependent. Equilibrium between bound and free enzyme was obtained within 90s, and Scatchard analysis indicated a high- and a low-affinity population of binding sites of approx. 1.24 × 10(4) sites/cell having a Kd of 1.4 × 10(-9) M and 7.2 × 10(4) sites/cell with a Kd of 2 × 10(-8) M respectively. Plasmin, bound to cell, was spontaneously released within 2 min, suggesting a rapid equilibrium. Chemical modification of the enzyme with phenylmethanesulphonyl fluoride or pyridoxal 5′-phosphate revealed that neither the active centre nor the heparin-binding site of plasmin was involved in the interaction with the endothelial cell. In terms of endothelial-cell receptors, the binding sites of cells for plasmin and thrombin were different: the two enzymes did not compete with each other, and the pretreatme...
Biochemical and Biophysical Research Communications, 1979
Cellular and Molecular Life Sciences, May 1, 1968
Nucleic Acids Research, 1974
A method is described for the preparation of p-aminophenyl oligo(dT)-Sepharose. This matrix has b... more A method is described for the preparation of p-aminophenyl oligo(dT)-Sepharose. This matrix has been used for the purification of polynucleotide phosphorylase from both E.coli and B.stearothermophilus. The effects of temperature and pH on the binding of the different enzymes to the matrix have been investigated. B.stearothermophilus isolated by affinity chromatography may be useful in selectively removing the polyA tract on the 3'-end of mRNA's.
Acta biochimica et biophysica Academiae Scientiarum Hungaricae, 1982
Le procede d'inhibition de la replication retrovirale consiste a manipuler la transferase ade... more Le procede d'inhibition de la replication retrovirale consiste a manipuler la transferase adenosine biphosphoribosyle enzymatique qui inhibite de maniere endogene la transcriptase retrovirale inverse. Le procede est utile pour la prevention et le traitement d'infections retrovirales, y compris HIV. L'inhibition de la replication virale est effectuee en administrant a un mammifere susceptible d'etre infecte ou infecte par un retrovirus des medicaments qui inhibent de maniere specifique la poly-ADP-ribosylation de la transferase d'adenosine biphosphoribosyle.
Blood, 1983
The interaction of human alpha-thrombin with mini-pig aortic endothelial cells was studied using ... more The interaction of human alpha-thrombin with mini-pig aortic endothelial cells was studied using 125I-labeled enzyme. Equilibrium between bound and free thrombin was attained within 1 min, and the Klotz-Hunston equations indicated two populations of binding sites. Approximately 30,000 sites/cell belonged to the high-affinity class with a Kd of about 3 x 10(-8) M. Modification of two lysine residues of thrombin with pyridoxal 5′-phosphate (PLP2-thrombin) destroyed the high- affinity binding and about three-fourths of the low-affinity bindings. When the lysine residue of thrombin involved in heparin binding was protected with heparin against chemical modification (PLP-thrombin), the modified enzyme remained similar to the native one with respect to cellular binding, with some loss of low-affinity binding only. Heparin, in a tenfold molar excess to enzyme, inhibited the binding of the native as well as the PLP-thrombin, whereas it did not influence the interaction between PLP2-thrombin...
Phytochemistry, 1972
A pyridoxal phosphate-dependent tryptophan decarboxylase has been purified 20-fold from seedlings... more A pyridoxal phosphate-dependent tryptophan decarboxylase has been purified 20-fold from seedlings of Phaluris tuberosa. The enzyme activity of the seedlings reached a maximum after 4 days. The enzyme activity is maximal at pH 7.6 and could be demonstrated either by the production of r4C-tryptamine from methylene 14C-L-tryptophan or of 14C02 from carboxyl-r4C-L-tryptophan. o-tryptophan was not a substrate but 5-hydroxy-14C-tryptophan was. The decarboxylation was inhibited at concentrations of 0.1 mM by a number of indole derivatives including N,N-dimethyltryptamine, one of the two major alkaloids produced by the plant. The second alkaloid, 5-methoxy-N,N-dimethyltryptamine did not inhibit at this concentration. A possible role of this enzyme in alkaloid biosynthesis is discussed.
PubMed, 1987
The proliferation stimulating or inhibitory effect of aortic endothelial cells on aortic smooth m... more The proliferation stimulating or inhibitory effect of aortic endothelial cells on aortic smooth muscle cells and vice versa was studied in cell culture by using a newly developed co-cultivation technique. The effect was registered as the incorporation of labelled thymidine detected by liquid scintillation counting. The experiments were performed in intra and interspecific combinations, using bovine, pig or rat aortic cell cultures. The endothelial cells stimulated smooth muscle cell proliferation in a species specific way. In interspecies combinations either inhibition or no effect was observed. The smooth muscle cells usually had an inhibitory effect on the proliferation of endothelial cells except for the pig smooth muscle cells which had a stimulatory effect in both inter and intraspecies combinations. Co-cultivation of smooth muscle or endothelial cells with cells of the same type resulted mostly in inhibition or no effect in both intra and interspecies combinations. This phenomenon was particularly conspicuous with endothelial cells. The different stimulatory and inhibitory effects produced by these cells may play an important role in the regulation of neovascularization and in reendothelization of denuded intimal areas.
PubMed, 1986
[U-14C]-glucose was incorporated non-enzymatically into the protein moiety of human low density l... more [U-14C]-glucose was incorporated non-enzymatically into the protein moiety of human low density lipoprotein. The incorporation was time and glucose concentration dependent. Investigated mini pig aortic endothelial cells showed that the glycosylated low density lipoprotein was bound, internalized and degraded significantly less than that of the native one measured with [125-I]-low density lipoprotein.
Differentiation, Dec 4, 2016
Fam3c, a cytokine-like protein, is a member of the Fam3 family (family with sequence similarity 3... more Fam3c, a cytokine-like protein, is a member of the Fam3 family (family with sequence similarity 3) and has been implicated to play a crucial role in Epithelial-to-mesenchymal transition (EMT) and subsequent metastasis during cancer progression. A few independent genome-wide association studies on different population cohorts predicted the gene locus of Fam3c to be associated with bone mineral density and fractures. In this study, we examined the role of Fam3c during osteoblast differentiation. Fam3c was found to be expressed during osteogenic differentiation of both primary bone marrow stromal cells and MC3T3-E1 pre-osteoblasts. In differentiating osteoblasts, knockdown of Fam3c increased alkaline phosphatase expression and activity whereas overexpression of Fam3c reduced it. Furthermore, overexpression of Fam3c caused reduction of Runx2 expression at both mRNA and protein levels. Fam3c was localized in the cytoplasm and it was not secreted outside the cell during osteoblast differentiation and therefore, may function intracellularly. Furthermore, Fam3c and TGF-β1 were found to regulate each other reciprocally. Our findings therefore suggest a functional role of Fam3c in the regulation of osteoblast differentiation. 2. Materials and methods 2.1. Cell culture MC3T3-E1 cells were cultured in α-MEM without ascorbic acid,
ADP-ribosyl transferase is a specific DNA-associating nuclear protein, whose enzymatic activity d... more ADP-ribosyl transferase is a specific DNA-associating nuclear protein, whose enzymatic activity depends on DNA-binding, probably to certain DNA sequences (1, 2). The study of molecular and biochemical mechanisms involving ADP-ribosyl transferase depends on experimentally efficient isolation of the enzyme. We present here a method that yields 8–9 mg of 95% homogeneous enzyme per kg starting material (calf thymus) within 3 days. In order to determine the polypeptide structure of ADP- ribosyl transferase, (with special reference to catalytically active and binding domains), we prepared stable polypeptides from ADP-ribosyl transferase by digestion with plasmin. We also report here on the partial sequencing of a polypeptide (36 kDa) that participates in the DNA-binding function of ADP- ribosyl transferase.
International Journal of Oncology, Dec 1, 1997
The GTPase activity of purified dimeric tubulin (alpha+beta) at 5 mu M was insensitive to methyl-... more The GTPase activity of purified dimeric tubulin (alpha+beta) at 5 mu M was insensitive to methyl-3,5-diiodo-4-(4'-methoxyphenoxy) benzoate (DIME), in contrast to nocodazole which activated GTPase. Cellular motility of MDA-MB-231 (human mammary cancer) cells migrating through 12-mu m pores was inhibited by DIME similar to nocodazole in a drug concentration-and DIME structure-dependent manner. An increase of cytoplasmic ATPase activity of DIME-treated cells without a decrease in ATP contents of intact cells suggests that DIME may also influence additional as yet unidentified ATP-dependent system(s) probably also involved in cell motility. These results show that DIME not only arrests cells in M phase but also inhibits cell motility in interphase. However the cellular mode of action of DIME is different from the action of other toxic tubulin-targeted drugs, despite the fact that DIME in a concentration-dependent manner disrupts microtubule structures in intact cells.
PubMed, 1990
To clarify whether some viruses could influence the different functions and membrane permeability... more To clarify whether some viruses could influence the different functions and membrane permeability of the aortic cells, we have examined in a model experiment the in vitro effect of the measles virus on the aortic endothelial and smooth muscle cells. The aortic cells infected with the virus failed to reveal gross cytopathic effect. Occasionally, however, syncytium formation and nuclear inclusions were observed. In infected endothelial cells lysosome containing viral nucleocapsids were seen. The early phase of measles virus replication inhibited the proliferation of endothelial cells of all species tested, while uniformly stimulated the replication of the smooth muscle cells relative to the control. In bovine aortic endothelial and smooth muscle cells the protein synthesis had been suppressed by the 4th to 6th hours postinfection. The results indicate that measles virus infection may be among the risk factors of atherosclerosis. It may damage endothelial cells by altering the cell membrane permeability and could induce proliferation of aortic smooth muscle cells.
Thrombosis and Haemostasis, 1983
SummaryHeparin forms a complex with human low density lipoprotein (LDL) in the presence of Ca2+. ... more SummaryHeparin forms a complex with human low density lipoprotein (LDL) in the presence of Ca2+. The complex is dissociable by 0.5 M NaCI. Thrombin and plasmin causes the dissociation of the LDL-heparin complex, whereas factor Xa does not. Heparin, complexed with LDL, retains its enhancing effect on the rate of thrombin and plasmin inactivation by antithrombin III. LDL isolated from the plasma of persons with different pathological conditions did not alter the rate of thrombin inactivation by antithrombin III either in the absence or in the presence of heparin. Heparin seems to maintain its biological functions when it is in a complex with LDL.
Molecular and Cellular Endocrinology, 2017
Fibroblast growth factors (FGF) and their receptors (FGFRs) regulate many developmental processes... more Fibroblast growth factors (FGF) and their receptors (FGFRs) regulate many developmental processes including differentiation of mesenchymal stromal cells (MSC). We developed two MSC lines capable of differentiating to osteoblasts and adipocytes and studied the role of FGFRs in this process. We identified FGFR2 and fibroblast growth factor receptor like-1 (FGFRL1) as possible actors in MSC differentiation with gene microarray and qRT-PCR. FGFR2 and FGFRL1 mRNA expression strongly increased during MSC differentiation to osteoblasts. FGF2 treatment, resulting in downregulation of FGFR2, or silencing FGFR2 expression with siRNAs inhibited osteoblast differentiation. During adipocyte differentiation expression of FGFR1 and FGFRL1 increased and was down-regulated by FGF2. FGFR1 knockdown inhibited adipocyte differentiation. Silencing FGFR2 and FGFR1 in MSCs was associated with decreased FGFRL1 expression in osteoblasts and adipocytes, respectively. Our results suggest that FGFR1 and FGFR2 regulate FGFRL1 expression. FGFRL1 may mediate or modulate FGFR regulation of MSC differentiation together with FGFR2 in osteoblastic and FGFR1 in adipocytic lineage.
Biochemical Journal, 1984
Interaction of human plasmin with a monolayer culture of mini-pig aortic endothelial cells was st... more Interaction of human plasmin with a monolayer culture of mini-pig aortic endothelial cells was studied by using the 125I-labelled enzyme. The binding of plasmin was time- and concentration-dependent. Equilibrium between bound and free enzyme was obtained within 90s, and Scatchard analysis indicated a high- and a low-affinity population of binding sites of approx. 1.24 × 10(4) sites/cell having a Kd of 1.4 × 10(-9) M and 7.2 × 10(4) sites/cell with a Kd of 2 × 10(-8) M respectively. Plasmin, bound to cell, was spontaneously released within 2 min, suggesting a rapid equilibrium. Chemical modification of the enzyme with phenylmethanesulphonyl fluoride or pyridoxal 5′-phosphate revealed that neither the active centre nor the heparin-binding site of plasmin was involved in the interaction with the endothelial cell. In terms of endothelial-cell receptors, the binding sites of cells for plasmin and thrombin were different: the two enzymes did not compete with each other, and the pretreatme...
Biochemical and Biophysical Research Communications, 1979
Cellular and Molecular Life Sciences, May 1, 1968
Nucleic Acids Research, 1974
A method is described for the preparation of p-aminophenyl oligo(dT)-Sepharose. This matrix has b... more A method is described for the preparation of p-aminophenyl oligo(dT)-Sepharose. This matrix has been used for the purification of polynucleotide phosphorylase from both E.coli and B.stearothermophilus. The effects of temperature and pH on the binding of the different enzymes to the matrix have been investigated. B.stearothermophilus isolated by affinity chromatography may be useful in selectively removing the polyA tract on the 3'-end of mRNA's.
Acta biochimica et biophysica Academiae Scientiarum Hungaricae, 1982
Le procede d'inhibition de la replication retrovirale consiste a manipuler la transferase ade... more Le procede d'inhibition de la replication retrovirale consiste a manipuler la transferase adenosine biphosphoribosyle enzymatique qui inhibite de maniere endogene la transcriptase retrovirale inverse. Le procede est utile pour la prevention et le traitement d'infections retrovirales, y compris HIV. L'inhibition de la replication virale est effectuee en administrant a un mammifere susceptible d'etre infecte ou infecte par un retrovirus des medicaments qui inhibent de maniere specifique la poly-ADP-ribosylation de la transferase d'adenosine biphosphoribosyle.
Blood, 1983
The interaction of human alpha-thrombin with mini-pig aortic endothelial cells was studied using ... more The interaction of human alpha-thrombin with mini-pig aortic endothelial cells was studied using 125I-labeled enzyme. Equilibrium between bound and free thrombin was attained within 1 min, and the Klotz-Hunston equations indicated two populations of binding sites. Approximately 30,000 sites/cell belonged to the high-affinity class with a Kd of about 3 x 10(-8) M. Modification of two lysine residues of thrombin with pyridoxal 5′-phosphate (PLP2-thrombin) destroyed the high- affinity binding and about three-fourths of the low-affinity bindings. When the lysine residue of thrombin involved in heparin binding was protected with heparin against chemical modification (PLP-thrombin), the modified enzyme remained similar to the native one with respect to cellular binding, with some loss of low-affinity binding only. Heparin, in a tenfold molar excess to enzyme, inhibited the binding of the native as well as the PLP-thrombin, whereas it did not influence the interaction between PLP2-thrombin...