Proteases influence colony aggregation behavior in Vibrio cholerae.

Autor:	Detomasi TC; Department of Chemistry, University of California, Berkeley, Berkeley, California, USA; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California, USA., Batka AE; Department of Chemistry, University of California, Berkeley, Berkeley, California, USA., Valastyan JS; Department of Molecular Biology, Princeton University, Princeton, New Jersey, USA; The Howard Hughes Medical Institute, Chevy Chase, Maryland, USA., Hydorn MA; Department of Chemistry, University of California, Berkeley, Berkeley, California, USA; Department of Microbiology and Immunology, College of Physicians and Surgeons, Columbia University, New York, New York, USA., Craik CS; Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California, USA., Bassler BL; Department of Molecular Biology, Princeton University, Princeton, New Jersey, USA; The Howard Hughes Medical Institute, Chevy Chase, Maryland, USA., Marletta MA; Department of Chemistry, University of California, Berkeley, Berkeley, California, USA; California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, California, USA; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA. Electronic address: marletta@berkeley.edu.
Jazyk:	angličtina
Zdroj:	The Journal of biological chemistry [J Biol Chem] 2023 Dec; Vol. 299 (12), pp. 105386. Date of Electronic Publication: 2023 Oct 26.
DOI:	10.1016/j.jbc.2023.105386
Abstrakt:	Aggregation behavior provides bacteria protection from harsh environments and threats to survival. Two uncharacterized proteases, LapX and Lap, are important for Vibrio cholerae liquid-based aggregation. Here, we determined that LapX is a serine protease with a preference for cleavage after glutamate and glutamine residues in the P1 position, which processes a physiologically based peptide substrate with a catalytic efficiency of 180 ± 80 M -1 s -1 . The activity with a LapX substrate identified by a multiplex substrate profiling by mass spectrometry screen was 590 ± 20 M -1 s -1 . Lap shares high sequence identity with an aminopeptidase (termed VpAP) from Vibrio proteolyticus and contains an inhibitory bacterial prepeptidase C-terminal domain that, when eliminated, increases catalytic efficiency on leucine p-nitroanilide nearly four-fold from 5.4 ± 4.1 × 10 4  M -1 s -1 to 20.3 ± 4.3 × 10 4  M -1 s -1 . We demonstrate that LapX processes Lap to its mature form and thus amplifies Lap activity. The increase is approximately eighteen-fold for full-length Lap (95.7 ± 5.6 × 10 4  M -1 s -1 ) and six-fold for Lap lacking the prepeptidase C-terminal domain (11.3 ± 1.9 × 10 5  M -1 s -1 ). In addition, substrate profiling reveals preferences for these two proteases that could inform in vivo function. Furthermore, purified LapX and Lap restore the timing of the V. cholerae aggregation program to a mutant lacking the lapX and lap genes. Both proteases must be present to restore WT timing, and thus they appear to act sequentially: LapX acts on Lap, and Lap acts on the substrate involved in aggregation. Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article. (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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