A conformational switch controls cell wall remodeling enzymes required for bacterial cell division
Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115, USA. Molecular Microbiology
(Impact Factor: 4.42).
06/2012; 85(4):768-81. DOI: 10.1111/j.1365-2958.2012.08138.x
Remodelling of the peptidoglycan (PG) exoskeleton is intimately tied to the growth and division of bacteria. Enzymes that hydrolyse PG are critical for these processes, but their activities must be tightly regulated to prevent the generation of lethal breaches in the PG matrix. Despite their importance, the mechanisms regulating PG hydrolase activity have remained elusive. Here we investigate the control of cell division hydrolases called amidases (AmiA, AmiB and AmiC) required for Escherichia coli cell division. Poorly regulated amiB mutants were isolated encoding lytic AmiB variants with elevated basal PG hydrolase activities in vitro. The structure of an AmiB orthologue was also solved, revealing that the active site of AmiB is occluded by a conserved alpha helix. Strikingly, most of the amino acid substitutions in the lytic AmiB variants mapped to this domain and are predicted to disrupt its interaction with the active site. Our results therefore support a model in which cell separation is stimulated by the reversible relief of amidase autoinhibition governed by conserved subcomplexes within the cytokinetic ring. Analogous conformational control mechanisms are likely to be part of a general strategy used to control PG hydrolases present within multienzyme PG-remodelling machines.
Available from: Felix Büttner
- "A flexible loop, not visible in the crystal structures, connects the insert to helix ␣3, which is significantly longer in AmiB and AmiC. LytM activators associated with septal ring factors allow a conformational shift of the inhibiting ␣-helix out of the binding site and therefore trigger amidase activity (Rocaboy et al., 2013; Yang et al., 2012). "
[Show abstract] [Hide abstract]
ABSTRACT: The molecular structure of matter defines its properties and function. This is especially true for biological macromolecules such as proteins, which participate in virtually all biochemical processes. A three dimensional structural model of a protein is thus essential for the detailed understanding of its physiological function and the characterization of essential properties such as ligand binding and reaction mechanism. X-ray crystallography is a well-established technique that has been used for many years, but it is still by far the most widely used method for structure determination. A particular strength of this technique is the elucidation of atomic details of molecular interactions, thus providing an invaluable tool for a multitude of scientific projects ranging from the structural classification of macromolecules over the validation of enzymatic mechanisms or the understanding of host-pathogen interactions to structure-guided drug design. In the first part of this review, we describe essential methodological and practical aspects of X-ray crystallography. We provide some pointers that should allow researchers without a background in structural biology to assess the overall quality and reliability of a crystal structure. To highlight its potential, we then survey the impact X-ray crystallography has had on advancing an understanding of a class of enzymes that modify the bacterial cell wall. A substantial number of different bacterial amidase structures have been solved, mostly by X-ray crystallography. Comparison of these structures highlights conserved as well as divergent features. In combination with functional analyses, structural information on these enzymes has therefore proven to be a valuable template not only for understanding their mechanism of catalysis, but also for targeted interference with substrate binding.
Available from: Richard A Daniel
- "The ABC transporter corresponds to a previously described cell division factor called FtsEX. It seems that the ATPase activity of the nucleotide-binding domain protein (FtsE) provokes a conformational change in the transmembrane component (FtsX) (Yang et al., 2012), which in turn activates the PG hydrolytic activity of the cognate autolysins: in Streptococcus pneumoniae this is a direct interaction with the putative PcsB autolysin (Sham et al., 2011); whereas in Escherichia coli activation works through an intermediate periplasmic protein called EnvC, and there are two regulated autolysins , AmiA and AmiB (Yang et al., 2011). "
[Show abstract] [Hide abstract]
ABSTRACT: Cell morphogenesis in most bacteria is governed by spatiotemporal growth regulation of the peptidoglycan cell wall layer. Much is known about peptidoglycan synthesis but regulation of its turnover by hydrolytic enzymes is much less well understood. B. subtilis has a multitude of such enzymes. Two of the best characterized are CwlO and LytE: cells lacking both enzymes have a lethal block in cell elongation. Here we show that activity of CwlO is regulated by an ABC transporter, FtsEX, which is required for cell elongation, unlike cell division as in Escherichia coli. Actin-like MreB proteins are thought to play a key role in orchestrating cell wall morphogenesis. B. subtilis has three MreB isologues with partially differentiated functions. We now show that the three MreB isologues have differential roles in regulation of the CwlO and LytE systems and that autolysins control different aspects of cell morphogenesis. The results add major autolytic activities to the growing list of functions controlled by MreB isologues in bacteria and provide new insights into the different specialised functions of essential cell wall autolysins.
Available from: Cécile Morlot
- "are missing conserved metal-binding and catalytic residues conserved in active LytM-like metallopeptidases (Uehara et al., 2010). We have thus proposed that the LytM factors promote a reversible conformational change in the amidases that stimulates the release of the inhibitory helix from the active site (Yang et al., 2012). Interestingly, the ability of EnvC to activate AmiA and AmiB in vivo is itself subject to the control of the ABC transporter-like complex composed of the divisome proteins FtsE and FtsX (Yang et al., 2011). "
[Show abstract] [Hide abstract]
ABSTRACT: Proteins with LytM (Peptidase_M23) domains are broadly distributed in bacteria and have been implicated in a variety of important processes, including cell division and cell-shape determination. Most LytM-like proteins that have been structurally and/or biochemically characterized are metallo-endopeptidases that cleave crosslinks in the peptidoglycan (PG) cell wall matrix. Notable exceptions are the Escherichia coli cell division proteins EnvC and NlpD. These LytM factors are not hydrolases themselves, but instead serve as activators that stimulate PG cleavage by target enzymes called amidases to promote cell separation. Here we report the structure of the LytM domain from EnvC, the first structure of a LytM factor implicated in the regulation of PG hydrolysis. As expected, the fold is highly similar to that of other LytM proteins. However, consistent with its role as a regulator, the active site region is degenerate and lacks a catalytic metal ion. Importantly, genetic analysis indicates that residues in and around this degenerate active site are critical for amidase activation in vivo and in vitro. Thus, in the regulatory LytM factors, the apparent substrate binding pocket conserved in active metallo-endopeptidases has been adapted to control PG hydrolysis by another set of enzymes.
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.