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Journal ArticleDOI

Endopeptidase penicillin-binding proteins 4 and 7 play auxiliary roles in determining uniform morphology of Escherichia coli

15 Dec 2004-Journal of Bacteriology (American Society for Microbiology)-Vol. 186, Iss: 24, pp 8326-8336
TL;DR: Overall cell shape may be determined by the existence or location of a specific type of peptide cross-link, with PBP 5 activity influencing how many cross- links are made and PBPs 4 and 7 acting as editing enzymes to remove inappropriate cross-links.
Abstract: The low-molecular-weight (LMW) penicillin-binding protein, PBP 5, plays a dominant role in determining the uniform cell shape of Escherichia coli. However, the physiological functions of six other LMW PBPs are unknown, even though the existence and enzymatic activities of four of these were established three decades ago. By applying fluorescence-activated cell sorting (FACS) to quantify the cellular dimensions of multiple PBP mutants, we found that the endopeptidases PBP 4 and PBP 7 also influence cell shape in concert with PBP 5. This is the first reported biological function for these two proteins. In addition, the combined loss of three dd-carboxypeptidases, PBPs 5 and 6 and DacD, also impaired cell shape. In contrast to previous reports based on visual inspection alone, FACS analysis revealed aberrant morphology in a mutant lacking only PBP 5, a phenotype not shared by any other strain lacking a single LMW PBP. PBP 5 removes the terminal d-alanine from pentapeptide side chains of muropeptide subunits, and pentapeptides act as donors for cross-linking adjacent side chains. As endopeptidases, PBPs 4 and 7 cleave cross-links in the cell wall. Therefore, overall cell shape may be determined by the existence or location of a specific type of peptide cross-link, with PBP 5 activity influencing how many cross-links are made and PBPs 4 and 7 acting as editing enzymes to remove inappropriate cross-links.
Citations
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Journal ArticleDOI
TL;DR: An overview of the content in PBPs of some bacteria is provided with an emphasis on comparing the biochemical properties of homologous PBPs (orthologues) belonging to different bacteria.
Abstract: Penicillin-binding proteins (PBPs) have been scrutinized for over 40 years. Recent structural information on PBPs together with the ongoing long-term biochemical experimental investigations, and results from more recent techniques such as protein localization by green fluorescent protein-fusion immunofluorescence or double-hybrid assay, have brought our understanding of the last stages of the peptidoglycan biosynthesis to an outstanding level that allows a broad outlook on the properties of these enzymes. Details are emerging regarding the interaction between the peptidoglycan-synthesizing PBPs and the peptidoglycan, their mesh net-like product that surrounds and protects bacteria. This review focuses on the detailed structure of PBPs and their implication in peptidoglycan synthesis, maturation and recycling. An overview of the content in PBPs of some bacteria is provided with an emphasis on comparing the biochemical properties of homologous PBPs (orthologues) belonging to different bacteria.

1,104 citations


Cites background from "Endopeptidase penicillin-binding pr..."

  • ...24 [9] Barrett DS, Chen L, Litterman NK & Walker S (2004) Expression and characterization of the 25 isolated glycosyltransferase module of Escherichia coli PBP1b....

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  • ...It has been suggested that both observed apoenzyme 22 structures may illustrate a conformational sampling of the ‘‘open’’ and ‘‘closed’’ forms that may 23 play a regulatory role in PBP1b catalytic activity....

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  • ...E. coli 21 possesses three class A PBPs. PBP1a and PBP1b are the major transpeptidases-transglycosylases....

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  • ...derivative of PBP1b (82-300) was also produced in the absence of detergent....

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  • ...For example, the fragments M46-D478 and M46-Q423 retain 23% 13 and 13% of the entire PBP1b activity respectively (Terrak, et al., 1999)....

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Journal ArticleDOI
TL;DR: The aim of this review is to spell out the physical, environmental, and biological forces that favor different bacterial morphologies and which, therefore, contribute to natural selection.
Abstract: Why do bacteria have shape? Is morphology valuable or just a trivial secondary characteristic? Why should bacteria have one shape instead of another? Three broad considerations suggest that bacterial shapes are not accidental but are biologically important: cells adopt uniform morphologies from among a wide variety of possibilities, some cells modify their shape as conditions demand, and morphology can be tracked through evolutionary lineages. All of these imply that shape is a selectable feature that aids survival. The aim of this review is to spell out the physical, environmental, and biological forces that favor different bacterial morphologies and which, therefore, contribute to natural selection. Specifically, cell shape is driven by eight general considerations: nutrient access, cell division and segregation, attachment to surfaces, passive dispersal, active motility, polar differentiation, the need to escape predators, and the advantages of cellular differentiation. Bacteria respond to these forces by performing a type of calculus, integrating over a number of environmental and behavioral factors to produce a size and shape that are optimal for the circumstances in which they live. Just as we are beginning to answer how bacteria create their shapes, it seems reasonable and essential that we expand our efforts to understand why they do so.

867 citations


Cites background from "Endopeptidase penicillin-binding pr..."

  • ...Cell shape is affected more and more as additional PBPs are removed (210, 229, 230), and the efficiency of biofilm formation decreases in step with increasingly aberrant shapes (86)....

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  • ...Deleting a single low-molecular-weight penicillin binding protein, PBP 5, causes cells to grow with nonuniform shape (210), which is enough to interfere with biofilm formation (86)....

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Journal ArticleDOI
TL;DR: The current view on the regulation of autolysins and on the role of cytoplasm hydrolases in peptidoglycan recycling and induction of beta-lactamase is reviewed.
Abstract: Most bacteria have multiple peptidoglycan hydrolases capable of cleaving covalent bonds in peptidoglycan sacculi or its fragments. An overview of the different classes of peptidoglycan hydrolases and their cleavage sites is provided. The physiological functions of these enzymes include the regulation of cell wall growth, the turnover of peptidoglycan during growth, the separation of daughter cells during cell division and autolysis. Specialized hydrolases enlarge the pores in the peptidoglycan for the assembly of large trans-envelope complexes (pili, flagella, secretion systems), or they specifically cleave peptidoglycan during sporulation or spore germination. Moreover, peptidoglycan hydrolases are involved in lysis phenomena such as fratricide or developmental lysis occurring in bacterial populations. We will also review the current view on the regulation of autolysins and on the role of cytoplasm hydrolases in peptidoglycan recycling and induction of β-lactamase.

785 citations


Cites background from "Endopeptidase penicillin-binding pr..."

  • ...Type-4 PBPs are generally considered as being indirectly involved in cell morphology (Meberg et al., 2004), in daughter cell separation (Priyadarshini et al....

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  • ...Type-4 PBPs are generally considered as being indirectly involved in cell morphology (Meberg et al., 2004), in daughter cell separation (Priyadarshini et al., 2006) and could be implicated in biofilm formation (Gallant et al., 2005) (see ‘Physiological functions of extracytoplasmic peptidoglycan…...

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Journal ArticleDOI
TL;DR: A model in which, at least for a subset of PBPs, the presence of substrate is a major factor in determining PBP localization is discussed, in both rod-shaped and coccoid cells.
Abstract: In order to maintain shape and withstand intracellular pressure, most bacteria are surrounded by a cell wall that consists mainly of the cross-linked polymer peptidoglycan (PG). The importance of PG for the maintenance of bacterial cell shape is underscored by the fact that, for various bacteria, several mutations affecting PG synthesis are associated with cell shape defects. In recent years, the application of fluorescence microscopy to the field of PG synthesis has led to an enormous increase in data on the relationship between cell wall synthesis and bacterial cell shape. First, a novel staining method enabled the visualization of PG precursor incorporation in live cells. Second, penicillin-binding proteins (PBPs), which mediate the final stages of PG synthesis, have been localized in various model organisms by means of immunofluorescence microscopy or green fluorescent protein fusions. In this review, we integrate the knowledge on the last stages of PG synthesis obtained in previous studies with the new data available on localization of PG synthesis and PBPs, in both rod-shaped and coccoid cells. We discuss a model in which, at least for a subset of PBPs, the presence of substrate is a major factor in determining PBP localization.

609 citations


Cites background or result from "Endopeptidase penicillin-binding pr..."

  • ...PBP5 is the major carboxypeptidase but is not specifically associated with cell division....

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  • ...H, and -4b; and PBP5 (169, 171) (Table 1)....

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  • ...However, a mutant with a knockout of the B. subtilis major carboxypeptidase, PBP5, displays normal morphology during exponential growth, although cells become progressively shorter after exponential growth (185)....

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  • ...LMW carboxypeptidase dacA (20) PBP5 Control of cell shape (117, 129, 130) dacC (20) PBP6 dacD (7) PBP6b...

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  • ...In E. coli strains defective for PBP5, a high frequency of branching is observed....

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Journal ArticleDOI
TL;DR: How a bacterium gains and maintains its shape, the challenges still confronting us and emerging strategies for answering difficult questions in this rapidly evolving field are discussed.
Abstract: Bacterial species have long been classified on the basis of their characteristic cell shapes. Despite intensive research, the molecular mechanisms underlying the generation and maintenance of bacterial cell shape remain largely unresolved. The field has recently taken an important step forward with the discovery that eukaryotic cytoskeletal proteins have homologues in bacteria that affect cell shape. Here, we discuss how a bacterium gains and maintains its shape, the challenges still confronting us and emerging strategies for answering difficult questions in this rapidly evolving field.

496 citations

References
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Book
01 Jan 1972
TL;DR: Molecular Genetics (Biology): An Overview | Sciencing Experimental in Molecular Genetics Experiments in molecular genetics (1972 edition) | Open ...
Abstract: Molecular Genetics (Biology): An Overview | Sciencing Experiments in Molecular Genetics Experiments in molecular genetics (1972 edition) | Open ... Experimental Molecular Genetics | Biology | MIT OpenCourseWare DNA experiments you can perform at home | SBS Science Experiments in molecular genetics Jeffrey H. Miller ... DNA and Molecular Genetics Experiments in Molecular Biology: Biochemical Applications ... Molecular Genetics Biology Experiment Please help ... Molecular genetics | biology | Britannica Molecular Genetic Experiment : Biology Lab 1793 Words ... Miller, J.H. (1972) Experiments in Molecular Genetics ... Griffith's experiment Wikipedia DNA as genetic material: Revisiting classic experiments ... Experiments in molecular genetics (Book, 1972) [WorldCat.org] Measuring βGalactosidase Activity in Bacteria: Cell ... Classic Experiments in

26,898 citations

Journal ArticleDOI
TL;DR: The tight regulation of the PBAD promoter is exploited to study the phenotypes of null mutations of essential genes and the use of pBAD vectors as an expression system is explored.
Abstract: We have constructed a series of plasmid vectors (pBAD vectors) containing the PBAD promoter of the araBAD (arabinose) operon and the gene encoding the positive and negative regulator of this promoter, araC. Using the phoA gene and phoA fusions to monitor expression in these vectors, we show that the ratio of induction/repression can be 1,200-fold, compared with 50-fold for PTAC-based vectors. phoA expression can be modulated over a wide range of inducer (arabinose) concentrations and reduced to extremely low levels by the presence of glucose, which represses expression. Also, the kinetics of induction and repression are very rapid and significantly affected by the ara allele in the host strain. Thus, the use of this system which can be efficiently and rapidly turned on and off allows the study of important aspects of bacterial physiology in a very simple manner and without changes of temperature. We have exploited the tight regulation of the PBAD promoter to study the phenotypes of null mutations of essential genes and explored the use of pBAD vectors as an expression system.

4,997 citations


Additional excerpts

  • ...of the arabinose promoter of pBAD18 (11)....

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Journal ArticleDOI
13 Jun 2003-Cell
TL;DR: A fluorescent derivative of the antibiotic vancomycin is used as a probe for nascent peptidoglycan synthesis in unfixed cells of various Gram-positive bacteria, providing insights into the diverse molecular strategies used by bacteria to control their cellular morphology, as well as suggesting ways in which these strategies may impact on growth rates and cell envelope structure.

751 citations

Journal ArticleDOI
TL;DR: The deltarecBCD::Plac-red kan replacement allele can be P1 transduced to other E. coli strains, making the hyper-Rec phenotype easily transferable.
Abstract: Replacement of Escherichia coli's RecBCD function with phage lambda's Red function generates a strain whose chromosome recombines with short linear DNA fragments at a greatly elevated rate. The rate is at least 70-fold higher than that exhibited by a recBC sbcBC or recD strain. The value of the system is highlighted by gene replacement with a PCR-generated DNA fragment. The deltarecBCD::Plac-red kan replacement allele can be P1 transduced to other E. coli strains, making the hyper-Rec phenotype easily transferable.

627 citations


"Endopeptidase penicillin-binding pr..." refers methods in this paper

  • ...The amplified PCR products were transferred by electroporation into E. coli KM-32 (21), where they were incorporated into the chromosome via homologous recombination mediated by the phage lambda recombination system as described previously (18, 21, 22)....

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  • ...KM-32 (21), where they were incorporated into the chromosome via homologous...

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Journal ArticleDOI
TL;DR: Evidence suggests that the Z ring is utilized by all prokaryotic organisms for division and may also be used by some eukaryotic organelles.
Abstract: Bacterial cell division occurs through the formation of an FtsZ ring (Z ring) at the site of division. The ring is composed of the tubulin-like FtsZ protein that has GTPase activity and the ability to polymerize in vitro. The Z ring is thought to function in vivo as a cytoskeletal element that is analogous to the contractile ring in many eukaryotic cells. Evidence suggests that the Z ring is utilized by all prokaryotic organisms for division and may also be used by some eukaryotic organelles. This review summarizes our present knowledge about the formation, function, and evolution of the Z ring in prokaryotic cell division.

479 citations


"Endopeptidase penicillin-binding pr..." refers background in this paper

  • ...The first is governed by a group of cytoskeleton proteins, including FtsZ, MreB, and Mbl, which polymerize as filamentous rings or helices on the inner face of the cytoplasmic membrane (3, 9, 15)....

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