Sortase-assembled pili in Corynebacterium diphtheriae are built using a latch mechanism

Journal Article · Mon Mar 15 00:00:00 EDT 2021 · Proceedings of the National Academy of Sciences of the United States of America

DOI:https://doi.org/10.1073/pnas.2019649118· OSTI ID:1847986

McConnell, Scott A.  ^[1]; McAllister, Rachel A. ^[1]; Amer, Brendan R.  ^[1]; Mahoney, Brendan J.  ^[1]; Sue, Christopher K. ^[1]; Chang, Chungyu  ^[2]; Ton-That, Hung ^[3]; Clubb, Robert T.  ^[4]

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1. Department of Chemistry and Biochemistry and the University of California, Los Angeles-US Department of Energy Institute of Genomics and Proteomics, University of California, Los Angeles, CA 90095,
2. Division of Oral Biology and Medicine, University of California, Los Angeles, CA 90095,
3. Division of Oral Biology and Medicine, University of California, Los Angeles, CA 90095,, Molecular Biology Institute, University of California, Los Angeles, CA 90095
4. Department of Chemistry and Biochemistry and the University of California, Los Angeles-US Department of Energy Institute of Genomics and Proteomics, University of California, Los Angeles, CA 90095,, Molecular Biology Institute, University of California, Los Angeles, CA 90095

Gram-positive bacteria assemble pili (fimbriae) on their surfaces to adhere to host tissues and to promote polymicrobial interactions. These hair-like structures, although very thin (1 to 5 nm), exhibit impressive tensile strengths because their protein components (pilins) are covalently crosslinked together via lysine–isopeptide bonds by pilus-specific sortase enzymes. While atomic structures of isolated pilins have been determined, how they are joined together by sortases and how these interpilin crosslinks stabilize pilus structure are poorly understood. Using a reconstituted pilus assembly system and hybrid structural biology methods, we elucidated the solution structure and dynamics of the crosslinked interface that is repeated to build the prototypical SpaA pilus from Corynebacterium diphtheriae. Here, we show that sortase-catalyzed introduction of a K190-T494 isopeptide bond between adjacent SpaA pilins causes them to form a rigid interface in which the LPLTG sorting signal is inserted into a large binding groove. Cellular and quantitative kinetic measurements of the crosslinking reaction shed light onto the mechanism of pilus biogenesis. We propose that the pilus-specific sortase in C. diphtheriae uses a latch mechanism to select K190 on SpaA for crosslinking in which the sorting signal is partially transferred from the enzyme to a binding groove in SpaA in order to facilitate catalysis. This process is facilitated by a conserved loop in SpaA, which after crosslinking forms a stabilizing latch that covers the K190-T494 isopeptide bond. General features of the structure and sortase-catalyzed assembly mechanism of the SpaA pilus are likely conserved in Gram-positive bacteria.

• Research Organization: Univ. of California, Los Angeles, CA (United States)
• Sponsoring Organization: USDOE Office of Science (SC), Biological and Environmental Research (BER); National Institutes of Health (NIH)
• Grant/Contract Number: FC03-02ER63421; AC02-05CH11231; AI52217; DE025015; DE017382; T32 GM007185; S10OD025073; S10OD016336; P30 GM124169; S10OD018483
• OSTI ID: 1847986
• Alternate ID(s): OSTI ID: 1833090
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America, Journal Name: Proceedings of the National Academy of Sciences of the United States of America Vol. 118 Journal Issue: 12; ISSN 0027-8424
• Publisher: Proceedings of the National Academy of Sciences
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
Corynebacterium diphtheriae
Pili
Sortase
Lysine isopeptide bond
Grampositive bacteria
NMR
Small angle x-ray scattering