Synthetic peptidoglycan substrates for penicillin-binding protein 5 of Gram-negative bacteria.
The major constituent of the bacterial cell wall, peptidoglycan, is comprised of repeating units of N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) with an appended peptide. Penicillin-binding proteins (PBPs) are involved in the final stages of bacterial cell wall assembly. Two activities for PBPs are the cross-linking of the cell wall, carried out by dd-transpeptidases, and the dd-peptidase activity, that removes the terminal d-Ala residue from peptidoglycan. The dd-peptidase activity moderates the extent of the cell wall cross-linking. There exists a balance between the two activities that is critical for the well-being of bacterial cells. We have cloned and purified PBP5 of Escherichia coli. The membrane anchor of this protein was removed, and the enzyme was obtained as a soluble protein. Two fragments of the polymeric cell wall of Gram-negative bacteria (compounds 5 and 6) were synthesized. These molecules served as substrates for PBP5. The products of the reactions of PBP5 and compounds 5 and 6 were isolated and were shown to be d-Ala and the fragments of the substrates minus the terminal d-Ala. The kinetic parameters for these enzymic reactions were evaluated. PBP5 would appear to have the potential for turnover of as many as 1.4 million peptidoglycan strands within a single doubling time (i.e., generation) of E. coli.
Hesek
Dusan
D
Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, Indiana 46556, USA.
Suvorov
Maxim
M
Morio
Ken-Ichiro
K
Lee
Mijoon
M
Brown
Stephen
S
Vakulenko
Sergei B
SB
Mobashery
Shahriar
S
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D006602:Hexosyltransferases / Q000235:genetics / Q000378:metabolism*; D002267:Muramoylpentapeptide Carboxypeptidase / Q000235:genetics / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular; D046915:Penicillin-Binding Proteins; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism; D010457:Peptidoglycan / Q000737:chemistry* / Q000378:metabolism*; D010458:Peptidyl Transferases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D046915:Penicillin-Binding Proteins; D010446:Peptide Fragments; D010457:Peptidoglycan; D011994:Recombinant Proteins; D010458:Peptidyl Transferases; D006602:Hexosyltransferases; D002267:Muramoylpentapeptide Carboxypeptidase