Bridging cell wall biosynthesis and bacterial morphogenesis

Matteï, Pierre-Jean; Neves, David; Dessen, Andréa

doi:10.1016/j.sbi.2010.09.014

Cited by 82 publications

(88 citation statements)

References 50 publications

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“…Cell wall synthesis presents another candidate process that could produce ATP-dependent fluctuations in bacteria. The peptidoglycan wall is constantly under construction by penicillin-binding proteins (PBPs), which catalyze the glycosyl transfer and transpeptidation reactions necessary to polymerize new cell wall (30,31). Although PBPs do not hydrolyze ATP directly, synthesis of their substrate, lipid II, involves ATPdependent reactions (32).…”

Section: Resultsmentioning

confidence: 99%

Nonthermal ATP-dependent fluctuations contribute to the in vivo motion of chromosomal loci

Weber

Spakowitz

Theriot

2012

Proc. Natl. Acad. Sci. U.S.A.

328

313

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Chromosomal loci jiggle in place between segregation events in prokaryotic cells and during interphase in eukaryotic nuclei. This motion seems random and is often attributed to Brownian motion. However, we show here that locus dynamics in live bacteria and yeast are sensitive to metabolic activity. When ATP synthesis is inhibited, the apparent diffusion coefficient decreases, whereas the subdiffusive scaling exponent remains constant. Furthermore, the magnitude of locus motion increases more steeply with temperature in untreated cells than in ATP-depleted cells. This "superthermal" response suggests that untreated cells have an additional source of molecular agitation, beyond thermal motion, that increases sharply with temperature. Such ATP-dependent fluctuations are likely mechanical, because the heat dissipated from metabolic processes is insufficient to account for the difference in locus motion between untreated and ATP-depleted cells. Our data indicate that ATP-dependent enzymatic activity, in addition to thermal fluctuations, contributes to the molecular agitation driving random (sub)diffusive motion in the living cell.T he cytoplasm is a crowded and dynamic medium, with molecules constantly jostling around and colliding with each other. This molecular motion is often attributed to Brownian motion, the random movement of suspended particles driven by thermal fluctuations of the solvent (1, 2). Classic Brownian motion theory assumes a system at thermal equilibrium. However, cells are far from equilibrium. They use the chemical energy of ATP (and GTP) to drive active biological processes, such as transport and metabolism.Recent work in eukaryotic cells demonstrates that biological activity generates nonthermal fluctuations of greater magnitude than thermal fluctuations (3-8). These active fluctuations can drive diffusive-like motion of molecules inside the cell, a phenomenon known as "active" diffusion (9, 10). In vitro experiments and analytical theory suggest that these active fluctuations are generated by the cytoskeletal molecular motor myosin (11-13). Thus, random molecular motion in vivo, at least in eukaryotic cytoplasm, may be due to active motor-driven forces in addition to passive thermal forces.Here we present evidence suggesting that ATP-dependent fluctuations contribute to the motion of chromosomal loci in bacterial and yeast cells. By modulating the temperature at which cells are observed, we were able to identify nonthermal forces that contribute to intracellular motion. Unlike active microrheology (7,8,11), temperature modulation presents a simple perturbation that can be applied to any experimental system to explore the physical processes underlying molecular motion in vivo. Our results suggest that "active" diffusion is not unique to systems containing eukaryotic cytoskeletal motors. This phenomenon may in fact be a general property of macromolecular motion in all living cells. Resultsis calculated to determine the subdiffusive scaling exponent α and the apparent diffusion coefficient D ap...

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Section: Resultsmentioning

confidence: 99%

Nonthermal ATP-dependent fluctuations contribute to the in vivo motion of chromosomal loci

Weber

Spakowitz

Theriot

2012

Proc. Natl. Acad. Sci. U.S.A.

328

313

View full text Add to dashboard Cite

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“…62 Although the details of the in vivo function of all of the enzymes mentioned above are not well understood, it is interesting to note that the soluble enzymes have no defined role in peptidoglycan synthesis and maintenance while the membrane-bound/associated enzymes have at least tentatively assigned roles. 2,48 One thus gets the impression that a number of these enzymes are present in solution in less than optimally specific and/or reactive conformations and that these cannot efficiently be induced into specific or reactive forms by small peptidoglycanmimetic peptide fragments. In some cases (LMMC), although not in others (LMMA), they can be so induced by tighter binding transition-state analogues.…”

Section: ■ Conclusionmentioning

confidence: 99%

Substrate Specificity of Low-Molecular Mass Bacterial dd-Peptidases

et al. 2011

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The bacterial DD-peptidases or penicillin-binding proteins (PBPs) catalyze the formation and regulation of cross-links in peptidoglycan biosynthesis. They are classified into two groups, the high-molecular mass (HMM) and lowmolecular mass (LMM) enzymes. The latter group, which is subdivided into classes A−C (LMMA, -B, and -C, respectively), is believed to catalyze DD-carboxypeptidase and endopeptidase reactions in vivo. To date, the specificity of their reactions with particular elements of peptidoglycan structure has not, in general, been defined. This paper describes the steady-state kinetics of hydrolysis of a series of specific peptidoglycan-mimetic peptides, representing various elements of stem peptide structure, catalyzed by a range of LMM PBPs (the LMMA enzymes, Escherichia coli PBP5, Neisseria gonorrhoeae PBP4, and Streptococcus pneumoniae PBP3, and the LMMC enzymes, the Actinomadura R39 DD-peptidase, Bacillus subtilis PBP4a, and N. gonorrhoeae PBP3). The R39 enzyme (LMMC), like the previously studied Streptomyces R61 DD-peptidase (LMMB), specifically and rapidly hydrolyzes stem peptide fragments with a free N-terminus. In accord with this result, the crystal structures of the R61 and R39 enzymes display a binding site specific to the stem peptide N-terminus. These are water-soluble enzymes, however, with no known specific function in vivo. On the other hand, soluble versions of the remaining enzymes of those noted above, all of which are likely to be membrane-bound and/or associated in vivo and have been assigned particular roles in cell wall biosynthesis and maintenance, show little or no specificity for peptides containing elements of peptidoglycan structure. Peptidoglycan-mimetic boronate transition-state analogues do inhibit these enzymes but display notable specificity only for the LMMC enzymes, where, unlike peptide substrates, they may be able to effectively induce a specific active site structure. The manner in which LMMA (and HMM) DD-peptidases achieve substrate specificity, both in vitro and in vivo, remains unknown. T he bacterial DD-peptidases catalyze the final steps of peptidoglycan (cell wall) biosynthesis 1,2 and are efficiently inhibited by β-lactam antibiotics.3 Resistance to these antibiotics continues to emerge, particularly from evolution of new β-lactamases, enzymes that catalyze hydrolytic destruction of β-lactams, but also by the evolution and dispersal of β-lactam-resistant DD-peptidases. and Neisseria gonorrhoeae. 10Many attempts have been made to develop new β-lactams effective against mutant 11,12 but, to date, these have not been completely successful. The ideal of a broadspectrum β-lactam not susceptible to loss of effectiveness through DD-peptidase mutation has been difficult to achieve. To a considerable extent, this is likely due to the largely trial and error approach to new β-lactams. The alternative approach of targeting the reaction center and the common and essential structural features of the DD-peptidase active site has not been pursued as vigorously. C...

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“…1). 14,22 High-molecular-mass (HMM) PBPs can either carry both GT and TP domains (class A), or catalyze only the TP reaction (class B). Low-molecular-mass (LMM) PBPs, on the other hand, can be either carboxypeptidases or endopeptidases, thus playing a role in the regulation of the level of PG cross-linking by cleaving peptide bonds within the stem peptide.…”

Section: Introductionmentioning

confidence: 99%