The cell wall envelope of Gram-positive bacteria can be thought of as a microbial organelle with anchored surface proteins that require specific signals and targeting mechanism for their assembly. Previous work identified sortases, enzymes that recognize short peptide sequences within sorting signals of secreted proteins. Upon cleavage of a protein substrate immediately following a conserved threonine residue, sortases form a thioester acyl intermediate between the C-terminal carboxyl group of the substrate and their active site cysteine residue. This intermediate is subsequently resolved by the nucleophilic attack of an amino group, which completes the transpeptidation reaction and restores the active site of sortase. Four classes of sortases are distinguished on the basis of sequence homology and enzyme function. One of these, sortase A, cleaves all surface proteins harboring LPXTG sorting signals and tethers its products to the amino group of lipid II, the precursor of cell wall biosynthesis. In contrast, sortase D cleaves only the sorting signals of pilin subunits. By accepting the side chain amino group of lysine (K) within the YPKN motif of pilin precursors, sortase D assembles pili on the surface of Gram-positive bacteria. Bacillus cereus pili are formed from two subunits, the tip protein (BcpB) and the major pilin (BcpA), which generates the pilus shaft. Sortases A and D collaborate on the topic of pilus formation;their assembled product is immobilized in the cell wall and extends 0.3-1.5

Public Health Relevance

Gram-positive bacteria cause many different human diseases and the morbidity and mortality of these infections, largely due to the development of antibiotic resistance traits, continues to increase in the United States. By studying the molecular mechanisms whereby Gram-positive bacteria promote the anchoring and surface display of proteins that enable the pathogenesis of human infections, we aim to increase knowledge about infectious processes. By developing inhibitors that prevent the assembly of proteins in the bacterial envelope we aim to develop antiinfectives that may be useful for the treatment of human infections.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI038897-18
Application #
8278552
Study Section
Prokaryotic Cell and Molecular Biology Study Section (PCMB)
Program Officer
Huntley, Clayton C
Project Start
1996-02-01
Project End
2014-05-31
Budget Start
2012-06-01
Budget End
2013-05-31
Support Year
18
Fiscal Year
2012
Total Cost
$378,250
Indirect Cost
$130,750
Name
University of Chicago
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
005421136
City
Chicago
State
IL
Country
United States
Zip Code
60637
Schneewind, Olaf; Missiakas, Dominique (2014) Genetic manipulation of Staphylococcus aureus. Curr Protoc Microbiol 32:Unit 9C.3.
Becker, Samuel; Frankel, Matthew B; Schneewind, Olaf et al. (2014) Release of protein A from the cell wall of Staphylococcus aureus. Proc Natl Acad Sci U S A 111:1574-9
Chan, Yvonne Gar-Yun; Kim, Hwan Keun; Schneewind, Olaf et al. (2014) The capsular polysaccharide of Staphylococcus aureus is attached to peptidoglycan by the LytR-CpsA-Psr (LCP) family of enzymes. J Biol Chem 289:15680-90
Zhang, Jie; Liu, Hongchuan; Zhu, Kongkai et al. (2014) Antiinfective therapy with a small molecule inhibitor of Staphylococcus aureus sortase. Proc Natl Acad Sci U S A 111:13517-22
Kim, Hwan Keun; Emolo, Carla; Missiakas, Dominique et al. (2014) A monoclonal antibody that recognizes the E domain of staphylococcal protein A. Vaccine 32:464-9
Deng, Fang-Kun; Zhang, Liang; Wang, Ya-Ting et al. (2014) Total chemical synthesis of the enzyme sortase A(?N59) with full catalytic activity. Angew Chem Int Ed Engl 53:4662-6
Kim, Hwan Keun; Missiakas, Dominique; Schneewind, Olaf (2014) Mouse models for infectious diseases caused by Staphylococcus aureus. J Immunol Methods 410:88-99
Cheng, Alice G; Missiakas, Dominique; Schneewind, Olaf (2014) The giant protein Ebh is a determinant of Staphylococcus aureus cell size and complement resistance. J Bacteriol 196:971-81
Thomer, Lena; Becker, Samuel; Emolo, Carla et al. (2014) N-acetylglucosaminylation of serine-aspartate repeat proteins promotes Staphylococcus aureus bloodstream infection. J Biol Chem 289:3478-86
Schneewind, Olaf; Missiakas, Dominique (2014) Sec-secretion and sortase-mediated anchoring of proteins in Gram-positive bacteria. Biochim Biophys Acta 1843:1687-97

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