Structure of the Bacillus anthracis Sortase A Enzyme Bound to Its Sorting Signal A FLEXIBLE AMINO-TERMINAL APPENDAGE MODULATES SUBSTRATE ACCESS

Albert H. Chan; Sung Wook Yi; Austen L. Terwilliger; Anthony W. Maresso; Michael E. Jung; Robert T. Clubb

doi:10.1074/jbc.M115.670984

Structure of the Bacillus anthracis Sortase A Enzyme Bound to Its Sorting Signal

A FLEXIBLE AMINO-TERMINAL APPENDAGE MODULATES SUBSTRATE ACCESS*

From the ^‡Department of Chemistry and Biochemistry,
^§UCLA-DOE Institute of Genomics and Proteomics, and
the ^¶Molecular Biology Institute, University of California, Los Angeles, California 90095 and
the ^‖Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas 77030

↵1 To whom correspondence should be addressed: Dept. of Chemistry and Biochemistry, University of California, Los Angeles, 602 Boyer Hall, Los Angeles, CA 90095. Tel.: 310-206-2334; Fax: 310-206-4749; E-mail: rclubb{at}mbi.ucla.edu.

Background: The Bacillus anthracis sortase A (^BaSrtA) enzyme attaches virulence factors to the cell surface.

Results: The structure of ^BaSrtA bound to a substrate analog reveals key amino acids involved in substrate recognition and catalysis.

Conclusion: ^BaSrtA modulates substrate access using a unique N-terminal appendage.

Significance: This research could facilitate the design of new anti-infective agents that work by disrupting surface protein display.

Abstract

The endospore forming bacterium Bacillus anthracis causes lethal anthrax disease in humans and animals. The ability of this pathogen to replicate within macrophages is dependent upon the display of bacterial surface proteins attached to the cell wall by the B. anthracis Sortase A (^BaSrtA) enzyme. Previously, we discovered that the class A ^BaSrtA sortase contains a unique N-terminal appendage that wraps around the body of the protein to contact the active site of the enzyme. To gain insight into its function, we determined the NMR structure of ^BaSrtA bound to a LPXTG sorting signal analog. The structure, combined with dynamics, kinetics, and whole cell protein display data suggest that the N terminus modulates substrate access to the enzyme. We propose that it may increase the efficiency of protein display by reducing the unproductive hydrolytic cleavage of enzyme-protein covalent intermediates that form during the cell wall anchoring reaction. Notably, a key active site loop (β7/β8 loop) undergoes a disordered to ordered transition upon binding the sorting signal, potentially facilitating recognition of lipid II.

Footnotes

↵* This work was supported, in whole or in part, by National Institutes of Health Grants AI52217 (to R. T. C. and M. E. J.), AI097167 and AI109465 (to A. W. M.), and T32 GM008496 (to A. H. C.) and Department of Energy Grant DE-FC-03-87ER60615. The authors declare that they have no conflicts of interest with the contents of this article.
The atomic coordinates and structure factors (code 2RUI) have been deposited in the Protein Data Bank (http://wwpdb.org/).
Chemical shift data have been deposited in the Biological Magnetic Resonance Data Bank under accession code 11570.
↵ This article contains supplemental Tables S1–S3.