Genetic and biochemical characterization of the cell wall hydrolase activity of the major secreted protein of Lactobacillus rhamnosus GG.

PloS One
Ingmar J J ClaesSarah Lebeer

Abstract

Lactobacillus rhamnosus GG (LGG) produces two major secreted proteins, designated here Msp1 (LGG_00324 or p75) and Msp2 (LGG_00031 or p40), which have been reported to promote the survival and growth of intestinal epithelial cells. Intriguingly, although each of these proteins shares homology with cell wall hydrolases, a physiological function that correlates with such an enzymatic activity remained to be substantiated in LGG. To investigate the bacterial function, we constructed knock-out mutants in the corresponding genes aiming to establish a genotype to phenotype relation. Microscopic examination of the msp1 mutant showed the presence of rather long and overly extended cell chains, which suggests that normal daughter cell separation is hampered. Subsequent observation of the LGG wild-type cells by immunofluorescence microscopy revealed that the Msp1 protein accumulates at the septum of exponential-phase cells. The cell wall hydrolyzing activity of the Msp1 protein was confirmed by zymogram analysis. Subsequent analysis by RP-HPLC and mass spectrometry of the digestion products of LGG peptidoglycan (PG) by Msp1 indicated that the Msp1 protein has D-glutamyl-L-lysyl endopeptidase activity. Immunofluorescence microscopy and th...Continue Reading

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Citations

Oct 29, 2013·Glycoconjugate Journal·Julia AnzengruberChristina Schäffer
Jun 3, 2014·BMC Genomics·Aminael Sánchez-RodríguezKathleen Marchal
Sep 5, 2014·Microbial Cell Factories·Marijke E Segers, Sarah Lebeer
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Jan 13, 2015·Applied and Environmental Microbiology·Cynthia E Vargas GarcíaSarah Lebeer
Jun 14, 2013·The Journal of Biological Chemistry·Thomas RolainPascal Hols
Mar 1, 2019·Scientific Reports·Camille Nina AllonsiusSarah Lebeer
Sep 19, 2017·Frontiers in Pharmacology·Giuseppe L BannaMassimo Libra
Apr 19, 2021·Trends in Microbiology·Mariya I PetrovaJessica A Ter Haar

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Methods Mentioned

BETA
electron microscopy
PCR
restriction digest

Software Mentioned

AxioVision

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