Nucleotide sequence of the pbpA gene and characteristics of the deduced amino acid sequence of penicillin‐binding protein 2 of Escherichia coli K12

Abstract

We have determined the nucleotide sequence of the pbpA gene encoding penicillin-binding protein (PBP) 2 of Escherichia coli. The coding region for PBP 2 was 1899 base pairs in length and was preceded by a possible promoter sequence and two open reading frames. The primary structure of PBP 2, deduced from the nucleotide sequence, comprised 633 amino acid residues. The relative molecular mass was calculated to be 70867. The deduced sequence agreed with the NH2-terminal sequence of PBP 2 purified from membranes, suggesting that PBP 2 has no signal peptide. The hydropathy profile suggested that the NH,-terminal hydrophobic region (a stretch of 25 non-ionic amino acids) may anchor PBP 2 in the cytoplasmic membrane as an ectoprotein. There were nine homologous segments in the amino acid sequence of PBP 2 when compared with PBP 3 of E. coli. The active-site serine residue of PBP 2 was predicted to be Ser-330. Around this putative active-site serine residue was found the conserved sequence of Ser-Xaa-Xaa-Lys, which has been identified in all of the other E. coli PBPs so far studied (PBPs lA, lB, 3, 5 and 6) and class A and class C B-lactamases. In the higher-molecular-mass PBPs 1 A, lB, 2 and 3, Ser-Xaa-Xaa-Lys-Pro was conserved. In the putative peptidoglycan transpeptidase domain there were six amino acid residues, which are common only in the PBPs of higher molecular mass.In Escherichia coli seven penicillin-binding proteins (PBPs) have been found in the cytoplasmic membrane [l -41. Among them, the higher-molecular-mass PBPs (PBP 1 A, PBP lB, PBP 2, and PBP 3 ) are important for completion of cell cycle of E. coli; PBP 1A and PBP 1 B are responsible for cell elongation [2, 3, 51, PBP 2 for the determination of rod shape of the cell [31,32], 3 [31, 331 and 5 [34] were obtained, and analyses of the peptides confirmed that the predicted active-site sequences were correct. The active-site serine regions of the higher-molecular-mass PBPs display a high sequence homology with each other [31].In this report we present the nucleotide sequence of the pbpA gene encoding PBP 2 of E. coli and the deduced amino acid sequence of this protein. These results prove the homology of the penicillin-binding sites of the higher-molecularmass PBPs of E. coli. Thus, the hypothesis of Tipper and Strominger [35] for the mechanism of the covalent binding of penicillin in all of the essential PBPs of E. coli has been verified.

Cloning and sequencing of the low-affinity penicillin-binding protein 3r-encoding gene of Enterococcus hirae S185: modular design and structural organization of the protein

“…The staphylococcal PBP2', which is responsible for acquired resistance to methicillin, is structurally related to the enterococcal PBP5 (7). The 633-aminoacid-residue PBP2 of E. coli is involved, together with the intrinsic membrane protein RodA (3,15,21), in the formation of the rod shape of the cell. Of the high-M, PBPs of known primary structure, the E. coli PBP2 shows the highest similarity with the low-affinity PBPs.…”

Bacterial Cell Wall Synthesis: New Insights from Localization Studies

“…The dibasic amino acid is often mesodiamoinopimelic acid (m-A 2 pm), which is present in most gram-negative bacteria and some gram-positive bacteria such as some Bacillus species, or L-lysine, which is present in most gram-positive bacteria. The most common stem peptide found in unprocessed PG in E. coli and B. subtilis is L-Ala (1) -DGlu (2) -m-A 2 pm (3) -D-Ala (4) -D-Ala (5) , with L-Ala (1) attached to the MurNac (57, 80, 175) (Fig. 2).…”

Cell-wall recycling and synthesis in Escherichia coli and Pseudomonas aeruginosa – their role in the development of resistance

“…High molecular mass penicillin-binding proteins (HMM PBPs) with a transglycosylase domain (PBP1A, PBP1B and PBP1C) facilitate the link between the MurNAc end of lipid II with the GlcNAc of the existing strand [45][46][47][48][49][50]. HMM PBPs with a transpeptidase domain (PBP1A, PBP1B, PBP1C, PBP2 and PBP3) catalyse the formation of cross-links between two peptide chains [47,[51][52][53][54] (Table 1). Cross-linking is composed of two steps wherein the terminal D-Ala-D-Ala of the peptide chain attached to MurNAc is cleaved with the release of one alanine [55].…”