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Biomedical subjects

L Rothfield

Publications and source records attributed to L Rothfield.

At least 19 recordsLinked to original sources
PubMed1999

Bacterial cell division.

Formation of the bacterial division septum is catalyzed by a number of essential proteins that assemble into a ring structure at the future division site. Assembly of proteins into the cytokinetic ring appears to occur in a hierarchial order that is initiated by the FtsZ protein, a structural and functional analog of eukaryotic tubulins. Placement of the division site at its correct location in Escherichia coli requires a division inhibitor (MinC), that is responsible for preventing septation at unwanted sites near the cell poles, and a topological specificity protein (MinE), that forms a ring at midcell and protects the midcell site from the division inhibitor. However, the mechanism responsible for identifying the position of the midcell site or the polar sites used for spore septum formation is still unclear. Regulation of the division process and its coordination with other cell cycle events, such as chromosome replication, are poorly understood. However, a protein has been identified in Caulobacter (CtrA) that regulates both the initiation of chromosome regulation and the transcription of ftsZ, and that may play an important role in the coordination process.

Cell Division↗
PubMed1998

Analysis of the length distribution of murein glycan strands in ftsZ and ftsI mutants of E. coli.

The chain length distribution of murein glycan strands was analyzed in wild-type cells and in cells in which preseptal and/or septal murein synthesis was prevented in ftsZ84 and ftsI36 mutants of E. coli. This revealed a significant change in glycan chain lengths in newly synthesized murein associated with inactivation of the ftsZ gene product but not with inactivation of the ftsI gene product. This is the first reported abnormality in murein biosynthesis associated with mutation of an essential cell division gene.

Bacterial Proteins↗
PubMed1998

High-affinity binding of hemimethylated oriC by Escherichia coli membranes is mediated by a multiprotein system that includes SeqA and a newly identified factor, SeqB.

The binding of hemimethylated oriC to Escherichia coli membranes has been implicated in the prevention of premature reinitiation at newly replicated chromosomal origins in a reaction that involves the SeqA protein. We describe the resolution of the membrane-associated oriC-binding activity into two fractions, both of which are required for the high-affinity binding of hemimethylated oriC. The active component in one fraction is identified as SeqA. The active component of the second fraction is a previously undescribed protein factor, SeqB. The reconstituted system reproduced the salient characteristics of the membrane-associated binding activity, suggesting that the membrane-associated oriC-binding machinery of E. coli is likely to be a multiprotein system that includes the SeqA and SeqB proteins.

Bacterial Outer Membrane Proteins↗
PubMed1998

The relationship between hetero-oligomer formation and function of the topological specificity domain of the Escherichia coli MinE protein.

MinE is an oligomeric protein that, in conjunction with other Min proteins, is required for the proper placement of the cell division site of Escherichia coli. We have examined the self-association properties of MinE by analytical ultracentrifugation and by studies of hetero-oligomer formation in non-denaturing polyacrylamide gels. The self-association properties of purified MinE predict that cytoplasmic MinE is likely to exist as a mixture of monomers and dimers. Consistent with this prediction, the C-terminal MinE22-88 fragment forms hetero-oligomers with MinE+ when the proteins are co-expressed. In contrast, the MinE36-88 fragment does not form MinE+/MinE36-88 hetero-oligomers, although MinE36-88 affects the topological specificity of septum placement as shown by its ability to induce minicell formation when co-expressed with MinE+ in wild-type cells. Therefore, hetero-oligomer formation is not necessary for the induction of minicelling by expression of MinE36-88 in wild-type cells. The interference with normal septal placement is ascribed to competition between MinE36-88 and the corresponding domain in the complete MinE protein for a component required for the topological specificity of septal placement.

Antibodies↗
PubMed1992

The essential bacterial cell-division protein FtsZ is a GTPase.

Cytokinesis defines the last stage in the division cycle, in which cell constriction leads to the formation of daughter cells. The biochemical mechanisms responsible for this process are poorly understood. In bacteria, the ftsZ gene product, FtsZ, is required for cell division, playing a prominent role in cytokinesis. The cellular concentration of FtsZ regulates the frequency of division and genetic studies have indicated that it is the target of several endogenous division inhibitors. At the time of onset of septal invagination, the FtsZ protein is recruited from the cytoplasm to the division site, where it assembles into a ring that remains associated with the leading edge of the invaginating septum until septation is completed. Here we report that FtsZ specifically binds and hydrolyses GTP. The reaction can be dissociated into a GTP-dependent activation stage that is markedly affected by the concentration of FtsZ, and a hydrolysis stage in which GTP is hydrolysed to GDP. The results indicate that GTP binding and hydrolysis are important in enabling FtsZ to support bacterial cytokinesis, either by facilitating the assembly of the FtsZ ring and/or by catalysing an essential step in the cytokinetic process itself.

Amino Acid Sequence↗
PubMed1992

Characterization of the Escherichia coli membrane domain responsible for binding oriC DNA.

It has previously been shown that hemimethylated DNA from the Escherichia coli replication origin (oriC) binds with high specificity to membrane fractions isolated from disrupted cells. In this article, the membrane localization of oriC-binding activity was studied by subjecting crude membrane preparations to successive cycles of sedimentation and flotation gradient analysis. This revealed that approximately two-thirds of the membrane-associated oriC-binding activity of the cell was not associated with the outer membrane fraction as previously suggested but was recovered instead in a unique membrane fraction (OCB1) whose buoyant density and protein profile differed from those of both inner and outer membranes. The specific activity of oriC binding in OCB1 was approximately fivefold higher than the activity of the isolated outer membrane peak. It is likely that membrane fraction OCB1 includes the membrane domain responsible for the binding of hemimethylated oriC to the cell envelope in intact cells.

Bacterial Outer Membrane Proteins↗
PubMed1992

Interactions of Escherichia coli membrane lipoproteins with the murein sacculus.

Bifunctional cross-linking reagents were used to identify cell envelope proteins that interacted with the murein sacculus. This revealed that a number of [3H]leucine-labeled proteins and [3H]palmitate-labeled lipoproteins were reproducibly cross-linked to the sacculus in plasmolyzed cells. The results suggested that most of the cell envelope lipoproteins, and not only the murein lipoprotein, mediate interactions between the murein sacculus and the inner and/or outer membrane of the cell.

Cross-Linking Reagents↗