TY - JOUR

T1 - Facile Production of the Pseudomonas aeruginosa Virulence Factor LasB in Escherichia coli for Structure-Based Drug Design

AU - Kolling, Dominik

AU - Haupenthal, Jörg

AU - Hirsch, Anna K.H.

AU - Koehnke, Jesko

N1 - Funding Information: 1 . We acknowledge the use of the ESRF beamlines ID30B and ID23‐1. We thank Simone Amann for the PA cultivation and supernatant preparation and Timo Risch for conducting the protein LC–MS measurements. We thank Dr. Andreas Kany for kindly providing compound . We thank Dr. Sally Shirran from the mass spectrometry and proteomics facility at St. Andrews University for conducting the tryptic LasB digesting and MS/MS measurements. Figure  1 2 and the table of contents graphic were prepared by using BioRender.com (2023). This work was supported by an ANR/BMBF grant (LasBAntiv, 16GW0346) to A.K.H.H. and J.K. Open Access funding enabled and organized by Projekt DEAL

PY - 2023/9/1

Y1 - 2023/9/1

N2 - The human pathogen Pseudomonas aeruginosa has a number of virulence factors at its disposal that play crucial roles in the progression of infection. LasB is one of the major virulence factors and exerts its effects through elastolytic and proteolytic activities aimed at dissolving connective tissue and inactivating host defense proteins. LasB is of great interest for the development of novel pathoblockers to temper the virulence, but access has thus far largely been limited to protein isolated from Pseudomonas cultures. Here, we describe a new protocol for high-level production of native LasB in Escherichia coli. We demonstrate that this facile approach is suitable for the production of mutant, thus far inaccessible LasB variants, and characterize the proteins biochemically and structurally. We expect that easy access to LasB will accelerate the development of inhibitors for this important virulence factor.

AB - The human pathogen Pseudomonas aeruginosa has a number of virulence factors at its disposal that play crucial roles in the progression of infection. LasB is one of the major virulence factors and exerts its effects through elastolytic and proteolytic activities aimed at dissolving connective tissue and inactivating host defense proteins. LasB is of great interest for the development of novel pathoblockers to temper the virulence, but access has thus far largely been limited to protein isolated from Pseudomonas cultures. Here, we describe a new protocol for high-level production of native LasB in Escherichia coli. We demonstrate that this facile approach is suitable for the production of mutant, thus far inaccessible LasB variants, and characterize the proteins biochemically and structurally. We expect that easy access to LasB will accelerate the development of inhibitors for this important virulence factor.

KW - anti-infective agents

KW - LasB

KW - pathoblockers

KW - virulence factors

UR - http://www.scopus.com/inward/record.url?scp=85166184173&partnerID=8YFLogxK

U2 - 10.1002/cbic.202300185

DO - 10.1002/cbic.202300185

M3 - Article

C2 - 37195753

AN - SCOPUS:85166184173

VL - 24

JO - CHEMBIOCHEM

JF - CHEMBIOCHEM

SN - 1439-4227

IS - 17

M1 - e202300185

ER -