Open AccessIn vitro reconstruction of the biosynthetic pathway of peptidoglycan cytoplasmic precursor in Pseudomonas aeruginosa
Author(s) -
El Zoeiby Ahmed,
Sanschagrin François,
Havugimana Pierre C.,
Garnier Alain,
Levesque Roger C.
Publication year - 2001
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2001.tb10761.x
Subject(s) - peptidoglycan , pseudomonas aeruginosa , in vitro , cytoplasm , microbiology and biotechnology , pseudomonadaceae , chemistry , pseudomonadales , bacteria , biology , biochemistry , cell wall , genetics
Bacterial peptidoglycan is the cell wall component responsible for maintaining cell integrity against osmotic pressure. Biosynthesis of the cytoplasmic precursor UDP‐ N ‐acetylmuramyl pentapeptide is catalyzed by the Mur enzymes. Genomic analysis of the three regions encoding Mur proteins was achieved. We have cloned and over‐expressed the murA , ‐ B , ‐ D , ‐ E and ‐ F genes of Pseudomonas aeruginosa in pET expression system by adding a His–Tag to the C‐termini of the proteins. Mur proteins were purified to homogeneity by a single chromatographic step on affinity nickel columns. Protein identities were verified through N‐terminal sequencing. Enzyme activity was proved by the identification of the pathway's final product.