Structure and activity of ChiX: a peptidoglycan hydrolase required for chitinase secretion by Serratia marcescens.
The Gram-negative bacterium Serratia marcescens secretes many proteins that are involved in extracellular chitin degradation. This so-called chitinolytic machinery includes three types of chitinase enzymes and a lytic polysaccharide monooxygenase. An operon has been identified in S. marcescens, chiWXYZ, that is thought to be involved in the secretion of the chitinolytic machinery. Genetic evidence points to the ChiX protein being a key player in the secretion mechanism, since deletion of the chiX gene in S. marcescens led to a mutant strain blocked for secretion of all members of the chitinolytic machinery. In this work, a detailed structural and biochemical characterisation of ChiX is presented. The high-resolution crystal structure of ChiX reveals the protein to be a member of the LAS family of peptidases. ChiX is shown to be a zinc-containing metalloenzyme, and in vitro assays demonstrate that ChiX is an l-Ala d-Glu endopeptidase that cleaves the cross-links in bacterial peptidoglycan. This catalytic activity is shown to be intimately linked with the secretion of the chitinolytic machinery, since substitution of the ChiX Asp-120 residue results in a variant protein that is both unable to digest peptidoglycan and cannot rescue the phenoytype of a chiX mutant strain.
['Owen|Richard A|RA|', 'Fyfe|Paul K|PK|', 'Lodge|Adam|A|', 'Biboy|Jacob|J|', 'Vollmer|Waldemar|W|', 'Hunter|William N|WN|', 'Sargent|Frank|F|0000-0002-6841-3624']
D020816:Amino Acid Motifs D001224:Aspartic Acid D001426:Bacterial Proteins D001665:Binding Sites D002686:Chitin D002688:Chitinases D003001:Cloning, Molecular D018360:Crystallography, X-Ray D004926:Escherichia coli D015870:Gene Expression D015964:Gene Expression Regulation, Bacterial D005822:Genetic Vectors D006868:Hydrolysis D008958:Models, Molecular D009238:N-Acetylmuramoyl-L-alanine Amidase D009876:Operon D010457:Peptidoglycan D011485:Protein Binding D000072756:Protein Conformation, alpha-Helical D000072757:Protein Conformation, beta-Strand D054730:Protein Interaction Domains and Motifs D011994:Recombinant Proteins D012706:Serratia marcescens D013379:Substrate Specificity D015032:Zinc
peptidoglycan hydrolase crystal structure anomalous dispersion bacterial chitinase secretion mutagenesis zinc enzyme