title: Structure-guided functional characterization of DUF1460 reveals a highly specific NlpC/P60 amidase family.
abstract: GlcNAc-1,6-anhydro-MurNAc-tetrapeptide is a major peptidoglycan degradation intermediate and a cytotoxin. It is generated by lytic transglycosylases and further degraded and recycled by various enzymes. We have identified and characterized a highly specific N-acetylmuramoyl-L-alanine amidase (AmiA) from Bacteroides uniformis, a member of the DUF1460 protein family, that hydrolyzes GlcNAc-1,6-anhydro-MurNAc-peptide into disaccharide and stem peptide. The high-resolution apo structure at 1.15 Å resolution shows that AmiA is related to NlpC/P60 γ-D-Glu-meso-diaminopimelic acid amidases and shares a common catalytic core and cysteine peptidase-like active site. AmiA has evolved structural adaptations that reconfigure the substrate recognition site. The preferred substrates for AmiA were predicted in silico based on structural and bioinformatics data, and subsequently were characterized experimentally. Further crystal structures of AmiA in complexes with GlcNAc-1,6-anhydro-MurNAc and GlcNAc have enabled us to elucidate substrate recognition and specificity. DUF1460 is highly conserved in structure and defines another amidase family.
authors.affiliation: Joint Center for Structural Genomics (JCSG); Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Menlo Park, CA 94025, USA.
authors.forename: Qingping
authors.initials: Q
authors.lastname: Xu
authors.affiliation: Laboratoire des Enveloppes Bactériennes et Antibiotiques, Université Paris-Sud, IBBMC, UMR 8619, Orsay F-91405, France; Centre National de la Recherche Scientifique (CNRS), Orsay F-91405, France. Electronic address: dominique.mengin-lecreulx@u-psud.fr.
authors.forename: Dominique
authors.initials: D
authors.lastname: Mengin-Lecreulx
authors.affiliation: Laboratoire des Enveloppes Bactériennes et Antibiotiques, Université Paris-Sud, IBBMC, UMR 8619, Orsay F-91405, France; Centre National de la Recherche Scientifique (CNRS), Orsay F-91405, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Patin
authors.affiliation: Joint Center for Structural Genomics (JCSG); Protein Sciences Department, Genomics Institute of the Novartis Research Foundation, San Diego, CA 92121, USA.
authors.forename: Joanna C
authors.initials: JC
authors.lastname: Grant
authors.affiliation: Joint Center for Structural Genomics (JCSG); Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Menlo Park, CA 94025, USA.
authors.forename: Hsiu-Ju
authors.initials: HJ
authors.lastname: Chiu
authors.affiliation: Joint Center for Structural Genomics (JCSG); Center for Research in Biological Systems, University of California, San Diego, La Jolla, CA 92093, USA; Program on Bioinformatics and Systems Biology, Sanford-Burnham Institute for Medical Research, La Jolla, CA 92037, USA.
authors.forename: Lukasz
authors.initials: L
authors.lastname: Jaroszewski
authors.affiliation: Joint Center for Structural Genomics (JCSG); Protein Sciences Department, Genomics Institute of the Novartis Research Foundation, San Diego, CA 92121, USA.
authors.forename: Mark W
authors.initials: MW
authors.lastname: Knuth
authors.affiliation: Joint Center for Structural Genomics (JCSG); Center for Research in Biological Systems, University of California, San Diego, La Jolla, CA 92093, USA; Program on Bioinformatics and Systems Biology, Sanford-Burnham Institute for Medical Research, La Jolla, CA 92037, USA.
authors.forename: Adam
authors.initials: A
authors.lastname: Godzik
authors.affiliation: Joint Center for Structural Genomics (JCSG); Protein Sciences Department, Genomics Institute of the Novartis Research Foundation, San Diego, CA 92121, USA; Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: Scott A
authors.initials: SA
authors.lastname: Lesley
authors.affiliation: Joint Center for Structural Genomics (JCSG); Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: Marc-André
authors.initials: MA
authors.lastname: Elsliger
authors.affiliation: Joint Center for Structural Genomics (JCSG); Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Menlo Park, CA 94025, USA.
authors.forename: Ashley M
authors.initials: AM
authors.lastname: Deacon
authors.affiliation: Joint Center for Structural Genomics (JCSG); Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA 92037, USA. Electronic address: wilson@scripps.edu.
authors.forename: Ian A
authors.initials: IA
authors.lastname: Wilson
mesh_terms: D001439:Bacteroides; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000378:metabolism*; D010457:Peptidoglycan / Q000378:metabolism*; D011487:Protein Conformation; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D010457:Peptidoglycan; D009238:N-Acetylmuramoyl-L-alanine Amidase