Jonathan Remis - Academia.edu (original) (raw)
Papers by Jonathan Remis
Microscopy and Microanalysis
http://journals.cambridge.org/abstract\_S1431927610057727
We have been observing BW Vul several nights per season in order to monitor the changes in its pe... more We have been observing BW Vul several nights per season in order to monitor the changes in its period. This paper describes some of the work carried out in collaboration with amateur astronomers.
PLoS ONE, 2014
The overwhelming majority of bacteria live in slime embedded microbial communities termed biofilm... more The overwhelming majority of bacteria live in slime embedded microbial communities termed biofilms, which are typically adherent to a surface. However, when several Staphylococcus epidermidis strains were cultivated in static liquid cultures, macroscopic aggregates were seen floating within the broth and also sedimented at the test tube bottom. Light-and electron microscopy revealed that early-stage aggregates consisted of bacteria and extracellular matrix, organized in sheetlike structures. Perpendicular under the sheets hung a network of periodically arranged, bacteria-associated strands. During the extended cultivation, the strands of a subpopulation of aggregates developed into cross-connected wall-like structures, in which aligned bacteria formed the walls. The resulting architecture had a compartmentalized appearance. In late-stage cultures, the wall-associated bacteria disintegrated so that, henceforth, the walls were made of the coalescing remnants of lysed bacteria, while the compartment-like organization remained intact. At the same time, the majority of strandcontaining aggregates with associated culturable bacteria continued to exist. These observations indicate that some strains of Staphylococcus epidermidis are able to build highly sophisticated structures, in which a subpopulation undergoes cell lysis, presumably to provide continued access to nutrients in a nutrient-limited environment, whilst maintaining structural integrity.
Science, 2004
The first structure of an ammonia channel from the Amt/MEP/Rh protein superfamily, determined to ... more The first structure of an ammonia channel from the Amt/MEP/Rh protein superfamily, determined to 1.35 angstrom resolution, shows it to be a channel that spans the membrane 11 times. Two structurally similar halves span the membrane with opposite polarity. Structures with and without ammonia or methyl ammonia show a vestibule that recruits NH 4 ϩ /NH 3 , a binding site for NH 4 ϩ , and a 20 angstrom-long hydrophobic channel that lowers the NH 4 ϩ pK a to below 6 and conducts NH 3 . Favorable interactions for NH 3 are seen within the channel and use conserved histidines. Reconstitution of AmtB into vesicles shows that AmtB conducts uncharged NH 3 .
Proceedings of the National Academy of Sciences, 2005
To explore the structural basis of the unique selectivity spectrum and conductance of the transme... more To explore the structural basis of the unique selectivity spectrum and conductance of the transmembrane channel protein AqpM from the archaeon Methanothermobacter marburgensis, we determined the structure of AqpM to 1.68-Å resolution by x-ray crystallography. The structure establishes AqpM as being in a unique subdivision between the two major subdivisions of aquaporins, the water-selective aquaporins, and the water-plus-glycerol-conducting aquaglyceroporins. In AqpM, isoleucine replaces a key histidine residue found in the lumen of water channels, which becomes a glycine residue in aquaglyceroporins. As a result of this and other side-chain substituents in the walls of the channel, the channel is intermediate in size and exhibits differentially tuned electrostatics when compared with the other subfamilies.
Journal of Bacteriology, 2009
Despite the fact that most bacteria grow in biofilms in natural and pathogenic ecosystems, very l... more Despite the fact that most bacteria grow in biofilms in natural and pathogenic ecosystems, very little is known about the ultrastructure of their component cells or about the details of their community architecture. We used high-pressure freezing and freeze-substitution to minimize the artifacts of chemical fixation, sample aggregation, and sample extraction. As a further innovation we have, for the first time in biofilm research, used electron tomography and three-dimensional (3D) visualization to better resolve the macromolecular 3D ultrastructure of a biofilm. This combination of superb specimen preparation and greatly improved resolution in the z axis has opened a window in studies of Myxococcus xanthus cell ultrastructure and biofilm community architecture. New structural information on the chromatin body, cytoplasmic organization, membrane apposition between adjacent cells, and structure and distribution of pili and vesicles in the biofilm matrix is presented. This paper is dedicated to the memory of Terry Beveridge, whose recent and untimely death robbed this field of one of its most perceptive and competent researchers.
Journal of Applied Crystallography, 2005
This paper describes the relatively simple modification of a light microscope to operate with ult... more This paper describes the relatively simple modification of a light microscope to operate with ultraviolet-based optics. Using a 280 nm UV illumination source, protein microcrystals are readily visualized in gels made from hydrated bilayers of phospholipids. Non-colored proteins stand out as clearly as colored proteins in this system, the imaging of which is based on UV absorption by tryptophan residues. In addition, protein crystals are easily distinguished from salt crystals. Artifacts from the lipid-based crystallization medium, which are frequently seen in brightfield microscopy, are greatly reduced when viewed in this UV-based microscope. laboratory notes J. Appl. Cryst. (2005). 38, 1031-1034 Christopher S. Lunde et al. UV microscopy at 280 nm 1033 Figure 3
Journal of Applied Crystallography, 2008
A simple and flexible system is described for in situ screening of microcrystals of membrane prot... more A simple and flexible system is described for in situ screening of microcrystals of membrane proteins that are grown within a connected-bilayer matrix formed by hydrated lipids. Using sheets of appropriate polymer materials to create a thin multiwell cassette, crystals can be evaluated by UV microscopy as well as by more conventional forms of light microscopy. Crystallization wells can be individually excised and mounted for diffraction screening on a synchrotron X-ray source. In addition, crystallization hit rates were significantly improved by employing a vapor diffusion approach rather than the batch crystallization method that is normally used with hydrated-lipid gels. laboratory notes J. Appl. Cryst. (2008). 41, 483-486 Christopher S. Lunde et al. Microcrystal screening 485
Environmental Microbiology, 2014
The social soil bacterium, Myxococcus xanthus, displays a variety of complex and highly coordinat... more The social soil bacterium, Myxococcus xanthus, displays a variety of complex and highly coordinated behaviours, including social motility, predatory rippling and fruiting body formation. Here we show that M. xanthus cells produce a network of outer membrane extensions in the form of outer membrane vesicle chains and membrane tubes that interconnect cells. We observed peritrichous display of vesicles and vesicle chains, and increased abundance in biofilms compared with planktonic cultures. By applying a range of imaging techniques, including three-dimensional (3D) focused ion beam scanning electron microscopy, we determined these structures to range between 30 and 60 nm in width and up to 5 μm in length. Purified vesicle chains consist of typical M. xanthus lipids, fucose, mannose, N-acetylglucosamine and N-acetylgalactoseamine carbohydrates and a small set of cargo protein. The protein content includes CglB and Tgl outer membrane proteins known to be transferable between cells in a contact-dependent manner. Most significantly, the 3D organization of cells within biofilms indicates that cells are connected via an extensive network of membrane extensions that may connect cells at the level of the periplasmic space. Such a network would allow the transfer of membrane proteins and other molecules between cells, and therefore could provide a mechanism for the coordination of social activities.
Current Opinion in Structural Biology, 2003
The aqua (glycero) porins conduct water (and glycerol) across cell membranes. The structure of th... more The aqua (glycero) porins conduct water (and glycerol) across cell membranes. The structure of these channels reveals a tripathic channel that supports a hydrophobic surface and, opposite to this, a line of eight hydrogen-bond acceptors and four hydrogen-bond donors. The eight carbonyls act as acceptors for water (or glycerol OH) molecules. The central water molecule in the channel is oriented to polarize hydrogen atoms outward from the center. This arrangement suggests how the structure prevents the potentially lethal conduction of protons across the membrane. The structure also suggests the mechanism behind the selectivity of aquaglyceroporins for glycerol, the basis for enantioselectivity among alditols, and the basis for the prevention of any leakage of the electrochemical gradient.
Numerous studies have helped characterize the stress response of the anaerobic sulfate reducer De... more Numerous studies have helped characterize the stress response of the anaerobic sulfate reducer Desulfovibrio vulgaris Hildenborough (DvH). Yet all of these techniques represent bulk analyses of cells grown mostly under liquid culture conditions in large reactors. Such results represent an average over a large variety of individual cellular responses, hence assuming a homogeneous distribution of physiological traits. Moreover, only recently
Frontiers in Microbiology, 2014
Myxococcus xanthus is a bacterial micro-predator known for hunting other microbes in a wolf pack-... more Myxococcus xanthus is a bacterial micro-predator known for hunting other microbes in a wolf pack-like manner. Outer membrane vesicles (OMVs) are produced in large quantities by M. xanthus and have a highly organized structure in the extracellular milieu, sometimes occurring in chains that link neighboring cells within a biofilm. OMVs may be a vehicle for mediating wolf pack activity by delivering hydrolytic enzymes and antibiotics aimed at killing prey microbes. Here, both the protein and small molecule cargo of the OMV and membrane fractions of M. xanthus were characterized and compared. Our analysis indicates a number of proteins that are OMV-specific or OMV-enriched, including several with putative hydrolytic function. Secondary metabolite profiling of OMVs identifies 16 molecules, many associated with antibiotic activities. Several hydrolytic enzyme homologs were identified, including the protein encoded by MXAN_3564 (mepA), an M36 protease homolog. Genetic disruption of mepA leads to a significant reduction in extracellular protease activity suggesting MepA is part of the long-predicted (yet to date undetermined) extracellular protease suite of M. xanthus.
Microscopy and Microanalysis
http://journals.cambridge.org/abstract\_S1431927610057727
We have been observing BW Vul several nights per season in order to monitor the changes in its pe... more We have been observing BW Vul several nights per season in order to monitor the changes in its period. This paper describes some of the work carried out in collaboration with amateur astronomers.
PLoS ONE, 2014
The overwhelming majority of bacteria live in slime embedded microbial communities termed biofilm... more The overwhelming majority of bacteria live in slime embedded microbial communities termed biofilms, which are typically adherent to a surface. However, when several Staphylococcus epidermidis strains were cultivated in static liquid cultures, macroscopic aggregates were seen floating within the broth and also sedimented at the test tube bottom. Light-and electron microscopy revealed that early-stage aggregates consisted of bacteria and extracellular matrix, organized in sheetlike structures. Perpendicular under the sheets hung a network of periodically arranged, bacteria-associated strands. During the extended cultivation, the strands of a subpopulation of aggregates developed into cross-connected wall-like structures, in which aligned bacteria formed the walls. The resulting architecture had a compartmentalized appearance. In late-stage cultures, the wall-associated bacteria disintegrated so that, henceforth, the walls were made of the coalescing remnants of lysed bacteria, while the compartment-like organization remained intact. At the same time, the majority of strandcontaining aggregates with associated culturable bacteria continued to exist. These observations indicate that some strains of Staphylococcus epidermidis are able to build highly sophisticated structures, in which a subpopulation undergoes cell lysis, presumably to provide continued access to nutrients in a nutrient-limited environment, whilst maintaining structural integrity.
Science, 2004
The first structure of an ammonia channel from the Amt/MEP/Rh protein superfamily, determined to ... more The first structure of an ammonia channel from the Amt/MEP/Rh protein superfamily, determined to 1.35 angstrom resolution, shows it to be a channel that spans the membrane 11 times. Two structurally similar halves span the membrane with opposite polarity. Structures with and without ammonia or methyl ammonia show a vestibule that recruits NH 4 ϩ /NH 3 , a binding site for NH 4 ϩ , and a 20 angstrom-long hydrophobic channel that lowers the NH 4 ϩ pK a to below 6 and conducts NH 3 . Favorable interactions for NH 3 are seen within the channel and use conserved histidines. Reconstitution of AmtB into vesicles shows that AmtB conducts uncharged NH 3 .
Proceedings of the National Academy of Sciences, 2005
To explore the structural basis of the unique selectivity spectrum and conductance of the transme... more To explore the structural basis of the unique selectivity spectrum and conductance of the transmembrane channel protein AqpM from the archaeon Methanothermobacter marburgensis, we determined the structure of AqpM to 1.68-Å resolution by x-ray crystallography. The structure establishes AqpM as being in a unique subdivision between the two major subdivisions of aquaporins, the water-selective aquaporins, and the water-plus-glycerol-conducting aquaglyceroporins. In AqpM, isoleucine replaces a key histidine residue found in the lumen of water channels, which becomes a glycine residue in aquaglyceroporins. As a result of this and other side-chain substituents in the walls of the channel, the channel is intermediate in size and exhibits differentially tuned electrostatics when compared with the other subfamilies.
Journal of Bacteriology, 2009
Despite the fact that most bacteria grow in biofilms in natural and pathogenic ecosystems, very l... more Despite the fact that most bacteria grow in biofilms in natural and pathogenic ecosystems, very little is known about the ultrastructure of their component cells or about the details of their community architecture. We used high-pressure freezing and freeze-substitution to minimize the artifacts of chemical fixation, sample aggregation, and sample extraction. As a further innovation we have, for the first time in biofilm research, used electron tomography and three-dimensional (3D) visualization to better resolve the macromolecular 3D ultrastructure of a biofilm. This combination of superb specimen preparation and greatly improved resolution in the z axis has opened a window in studies of Myxococcus xanthus cell ultrastructure and biofilm community architecture. New structural information on the chromatin body, cytoplasmic organization, membrane apposition between adjacent cells, and structure and distribution of pili and vesicles in the biofilm matrix is presented. This paper is dedicated to the memory of Terry Beveridge, whose recent and untimely death robbed this field of one of its most perceptive and competent researchers.
Journal of Applied Crystallography, 2005
This paper describes the relatively simple modification of a light microscope to operate with ult... more This paper describes the relatively simple modification of a light microscope to operate with ultraviolet-based optics. Using a 280 nm UV illumination source, protein microcrystals are readily visualized in gels made from hydrated bilayers of phospholipids. Non-colored proteins stand out as clearly as colored proteins in this system, the imaging of which is based on UV absorption by tryptophan residues. In addition, protein crystals are easily distinguished from salt crystals. Artifacts from the lipid-based crystallization medium, which are frequently seen in brightfield microscopy, are greatly reduced when viewed in this UV-based microscope. laboratory notes J. Appl. Cryst. (2005). 38, 1031-1034 Christopher S. Lunde et al. UV microscopy at 280 nm 1033 Figure 3
Journal of Applied Crystallography, 2008
A simple and flexible system is described for in situ screening of microcrystals of membrane prot... more A simple and flexible system is described for in situ screening of microcrystals of membrane proteins that are grown within a connected-bilayer matrix formed by hydrated lipids. Using sheets of appropriate polymer materials to create a thin multiwell cassette, crystals can be evaluated by UV microscopy as well as by more conventional forms of light microscopy. Crystallization wells can be individually excised and mounted for diffraction screening on a synchrotron X-ray source. In addition, crystallization hit rates were significantly improved by employing a vapor diffusion approach rather than the batch crystallization method that is normally used with hydrated-lipid gels. laboratory notes J. Appl. Cryst. (2008). 41, 483-486 Christopher S. Lunde et al. Microcrystal screening 485
Environmental Microbiology, 2014
The social soil bacterium, Myxococcus xanthus, displays a variety of complex and highly coordinat... more The social soil bacterium, Myxococcus xanthus, displays a variety of complex and highly coordinated behaviours, including social motility, predatory rippling and fruiting body formation. Here we show that M. xanthus cells produce a network of outer membrane extensions in the form of outer membrane vesicle chains and membrane tubes that interconnect cells. We observed peritrichous display of vesicles and vesicle chains, and increased abundance in biofilms compared with planktonic cultures. By applying a range of imaging techniques, including three-dimensional (3D) focused ion beam scanning electron microscopy, we determined these structures to range between 30 and 60 nm in width and up to 5 μm in length. Purified vesicle chains consist of typical M. xanthus lipids, fucose, mannose, N-acetylglucosamine and N-acetylgalactoseamine carbohydrates and a small set of cargo protein. The protein content includes CglB and Tgl outer membrane proteins known to be transferable between cells in a contact-dependent manner. Most significantly, the 3D organization of cells within biofilms indicates that cells are connected via an extensive network of membrane extensions that may connect cells at the level of the periplasmic space. Such a network would allow the transfer of membrane proteins and other molecules between cells, and therefore could provide a mechanism for the coordination of social activities.
Current Opinion in Structural Biology, 2003
The aqua (glycero) porins conduct water (and glycerol) across cell membranes. The structure of th... more The aqua (glycero) porins conduct water (and glycerol) across cell membranes. The structure of these channels reveals a tripathic channel that supports a hydrophobic surface and, opposite to this, a line of eight hydrogen-bond acceptors and four hydrogen-bond donors. The eight carbonyls act as acceptors for water (or glycerol OH) molecules. The central water molecule in the channel is oriented to polarize hydrogen atoms outward from the center. This arrangement suggests how the structure prevents the potentially lethal conduction of protons across the membrane. The structure also suggests the mechanism behind the selectivity of aquaglyceroporins for glycerol, the basis for enantioselectivity among alditols, and the basis for the prevention of any leakage of the electrochemical gradient.
Numerous studies have helped characterize the stress response of the anaerobic sulfate reducer De... more Numerous studies have helped characterize the stress response of the anaerobic sulfate reducer Desulfovibrio vulgaris Hildenborough (DvH). Yet all of these techniques represent bulk analyses of cells grown mostly under liquid culture conditions in large reactors. Such results represent an average over a large variety of individual cellular responses, hence assuming a homogeneous distribution of physiological traits. Moreover, only recently
Frontiers in Microbiology, 2014
Myxococcus xanthus is a bacterial micro-predator known for hunting other microbes in a wolf pack-... more Myxococcus xanthus is a bacterial micro-predator known for hunting other microbes in a wolf pack-like manner. Outer membrane vesicles (OMVs) are produced in large quantities by M. xanthus and have a highly organized structure in the extracellular milieu, sometimes occurring in chains that link neighboring cells within a biofilm. OMVs may be a vehicle for mediating wolf pack activity by delivering hydrolytic enzymes and antibiotics aimed at killing prey microbes. Here, both the protein and small molecule cargo of the OMV and membrane fractions of M. xanthus were characterized and compared. Our analysis indicates a number of proteins that are OMV-specific or OMV-enriched, including several with putative hydrolytic function. Secondary metabolite profiling of OMVs identifies 16 molecules, many associated with antibiotic activities. Several hydrolytic enzyme homologs were identified, including the protein encoded by MXAN_3564 (mepA), an M36 protease homolog. Genetic disruption of mepA leads to a significant reduction in extracellular protease activity suggesting MepA is part of the long-predicted (yet to date undetermined) extracellular protease suite of M. xanthus.