scholarly journals The Enterococcus hirae R40 penicillin-binding protein 5 and the methicillin-resistant Staphylococcus aureus penicillin-binding protein 2′ are similar

1991 ◽  
Vol 280 (2) ◽  
pp. 463-469 ◽  
Author(s):  
A el Kharroubi ◽  
P Jacques ◽  
G Piras ◽  
J Van Beeumen ◽  
J Coyette ◽  
...  
Keyword(s):  
Amino Acid ◽  
Binding Protein ◽  
Water Soluble ◽  
Penicillin Binding Protein ◽  
Enterococcus Hirae ◽  
Methicillin Resistant ◽  
Class B ◽  
Membrane Bound ◽  
Polymerase Chain ◽  
Terminal Domains

The penicillin-resistant Enterococcus hirae R40 has a typical profile of membrane-bound penicillin-binding proteins (PBPs) except that the 71 kDa PBP5 of low penicillin affinity represents about 50% of all the PBPs present. Water-soluble tryptic-digest peptides were selectively produced from PBP5, their N-terminal regions were sequenced and synthetic oligonucleotides were used as primers to generate a 476 bp DNA fragment by polymerase chain reaction. On the basis of these data, the PBP5-encoding gene was cloned in Escherichia coli by using pBR322 as vector. The gene, included in a 7.1 kb insert, had the information for a 678-amino acid-residue protein. PBP5 shows similarity, in the primary structure, with the high-molecular-mass PBPs of class B. In particular, amino acid alignment of the enterococcal PBP5 and the methicillin-resistant staphylococcal PBP2′ generates scores that are 30, for the N-terminal domains, and 53, for the C-terminal domains, standard deviations above that expected for a run of 20 randomized pairs of proteins having the same amino acid compositions as the two proteins under consideration.

scholarly journals Active-site and membrane topology of the DD-peptidase/penicillin-binding protein no. 6 of Enterococcus hirae (Streptococcus faecium) A.T.C.C. 9790

1989 ◽  
Vol 262 (2) ◽  
pp. 457-462 ◽  
Author(s):  
A el Kharroubi ◽  
G Piras ◽  
P Jacques ◽  
I Szabo ◽  
J Van Beeumen ◽  
...  
Keyword(s):  
Active Site ◽  
Binding Capacity ◽  
Binding Protein ◽  
Membrane Topology ◽  
Penicillin Binding Protein ◽  
Enterococcus Hirae ◽  
Serine Residue ◽  
Membrane Bound ◽  
High Degree ◽  
Terminal Appendage

The membrane-bound 43,000-Mr penicillin-binding protein no. 6 (PBP6) of Enterococcus hirae consists of a 30,000-Mr DD-peptidase/penicillin-binding domain and a approximately 130-residue C-terminal appendage. Removal of this appendage by trypsin proteolysis has no marked effect on the catalytic activity and penicillin-binding capacity of the PBP. Anchorage of the PBP in the membrane appears to be mediated by a short 15-20-residue stretch at the C-terminal end of the appendage. The sequence of the 50-residue N-terminal region of the PBP shows high degree of homology with the sequences of the corresponding regions of the PBPs5 of Escherichia coli and Bacillus subtilis. On this basis the active-site serine residue occurs at position 35 in the enterococcal PBP.

scholarly journals Secretion by overexpression and purification of the water-soluble Streptomyces K15 dd-transpeptidase/penicillin-binding protein

1992 ◽  
Vol 288 (1) ◽  
pp. 87-91 ◽  
Author(s):  
P Palomeque-Messia ◽  
V Quittre ◽  
M Leyh-Bouille ◽  
M Nguyen-Distèche ◽  
C J L Gershater ◽  
...  
Keyword(s):  
Culture Supernatant ◽  
Binding Capacity ◽  
Binding Protein ◽  
Water Soluble ◽  
Appreciable Amount ◽  
Soluble Enzyme ◽  
Penicillin Binding Protein ◽  
Wild Type ◽  
Wild Type Enzyme ◽  
Membrane Bound

Though synthesized with a cleavable signal peptide and devoid of membrane anchors, the 262-amino-acid-residue Streptomyces K15 DD-transpeptidase/penicillin-binding protein is membrane-bound. Overexpression in Streptomyces lividans resulted in the export of an appreciable amount of the synthesized protein (4 mg/litre of culture supernatant). The water-soluble enzyme was purified close to protein homogeneity with a yield of 75%. It requires the presence of 0.5 M-NaCl to remain soluble. It is indistinguishable from the detergent-extract wild-type enzyme with respect to molecular mass, thermostability, transpeptidase activity and penicillin-binding capacity.

scholarly journals Engineering and overexpression of periplasmic forms of the penicillin-binding protein 3 of Escherichia coli

1994 ◽  
Vol 298 (1) ◽  
pp. 189-195 ◽  
Author(s):  
C Fraipont ◽  
M Adam ◽  
M Nguyen-Distèche ◽  
W Keck ◽  
J Van Beeumen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Binding Protein ◽  
Molar Ratio ◽  
Amino Acid Residues ◽  
Penicillin Binding Protein ◽  
Operator Expression ◽  
Membrane Bound ◽  
Terminal Amino ◽  
Laci Repressor

Replacement of the 36 and 56 N-terminal amino acid residues of the 588-amino-acid-residue membrane-bound penicillin-binding protein 3 (PBP3) of Escherichia coli by the OmpA signal peptide allows export of F37-V577 PBP3 and G57-V577 PBP3 respectively into the periplasm. The modified ftsI genes were placed under the control of the fused lpp promoter and lac promoter/operator; expression of the truncated PBP3s was optimized by varying the copy number of the recombinant plasmids and the amount of LacI repressor, and export was facilitated by increasing the SecB content of the producing strain. The periplasmic PBP3s (yield 8 mg/l of culture) were purified to 70% protein homogeneity. They require the presence of 0.25 M NaCl to remain soluble. Like the membrane-bound PBP3, they undergo processing by elimination of the C-terminal decapeptide I578-S588, they bind penicillin in a 1:1 molar ratio and they catalyse hydrolysis and aminolysis of acyclic thioesters that are analogues of penicillin. The membrane-anchor-free PBP3s have ragged N-termini. The G57-V577 PBP3, however, is less prone to proteolytic degradation than the F37-V577 PBP3.

scholarly journals Ceftaroline Resistance by Clone-Specific Polymorphism in Penicillin-Binding Protein 2a of Methicillin-Resistant Staphylococcus aureus

2018 ◽  
Vol 62 (9) ◽  
Author(s):  
Hyukmin Lee ◽  
Eun-Jeong Yoon ◽  
Dokyun Kim ◽  
Jung Wook Kim ◽  
Kwang-Jun Lee ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Amino Acid ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Binding Protein ◽  
Amino Acid Substitutions ◽  
Penicillin Binding Protein ◽  
Methicillin Resistant ◽  
Content Type ◽  
Cefoxitin Disk ◽  
Cefoxitin Disk Diffusion

ABSTRACT A total of 281 nonduplicated Staphylococcus aureus blood isolates were collected from January to May 2017 from eight hospitals in South Korea to investigate the epidemiological traits of ceftaroline resistance in methicillin-resistant S. aureus (MRSA). Cefoxitin-disk diffusion tests and the mecA gene PCR revealed that 56.6% (159/281) of the S. aureus isolates were MRSA, and most belonged to ST5 (50.3%, 80/281) and ST72 (41.5%, 66/281). Of the MRSA isolates, 44.0% (70/159) were nonsusceptible to ceftaroline (MIC ≥ 2 mg/liter), whereas all of the methicillin-susceptible S. aureus isolates were susceptible to the drug. Eight amino acid substitutions in penicillin-binding protein 2a (PBP2a), including four (L357I, E447K, I563T, and S649A) in the penicillin-binding domain (PBD) and four (N104K, V117I, N146K, and A228V) in the non-PBD (nPBD) of PBP2a, were associated with ceftaroline resistance. The accumulation of substitutions in PBP2a resulted in the elevation of ceftaroline MICs: one substitution at 1 to 2 mg/liter, two or three substitutions at 2 to 4 mg/liter, and five substitutions at 4 or 16 mg/liter. Ceftaroline resistance in MRSA might be the result of clone-specific PBP2a polymorphism, along with substitutions both in PBD and nPBD, and the elevated ceftaroline MICs were associated with the substitution sites and accumulation of substitutions.

scholarly journals Increasing Ceftriaxone Resistance and Multiple Alterations of Penicillin-Binding Proteins among Penicillin-Resistant Streptococcus pneumoniae Isolates in Taiwan

2007 ◽  
Vol 51 (9) ◽  
pp. 3404-3406 ◽  
Author(s):  
Cheng-Hsun Chiu ◽  
Lin-Hui Su ◽  
Yhu-Chering Huang ◽  
Jui-Chia Lai ◽  
Hsiu-Ling Chen ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Amino Acid ◽  
Binding Proteins ◽  
Binding Protein ◽  
Penicillin Binding Protein ◽  
Penicillin Binding Proteins

ABSTRACT The rate of nonsusceptibility of penicillin-resistant Streptococcus pneumoniae strains to ceftriaxone increased significantly in Taiwan in 2005. Approximately 90% of the ceftriaxone-nonsusceptible isolates were found to be of four major serotypes (serotypes 6B, 14, 19F, and 23F). Seven amino acid alterations in the penicillin-binding protein 2B transpeptidase-encoding region specifically contributed to the resistance.

scholarly journals Amino acid sequence of the penicillin-binding protein/dd-peptidase of Streptomyces K15. Predicted secondary structures of the low Mr penicillin-binding proteins of class A

1991 ◽  
Vol 279 (1) ◽  
pp. 223-230 ◽  
Author(s):  
P Palomeque-Messia ◽  
S Englebert ◽  
M Leyh-Bouille ◽  
M Nguyen-Distèche ◽  
C Duez ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Residue ◽  
Active Sites ◽  
Binding Protein ◽  
Secondary Structures ◽  
Peptide Sequence ◽  
Penicillin Binding Protein ◽  
Signal Peptide Sequence ◽  
Beta Lactamases ◽  
Class A

The low-Mr penicillin-binding protein (PBP)/DD-transpeptidase of Streptomyces K15 is synthesized in the form of a 291-amino acid-residue precursor possessing a cleavable 29-amino acid-residue signal peptide. Sequence-similarity searches and hydrophobic-cluster analysis show that the Streptomyces K15 enzyme, the Escherichia coli PBPs/DD-carboxy-peptidases 5 and 6, the Bacillus subtilis PBP/DD-carboxypeptidase 5 and the spoIIA product (a putative PBP involved in the sporulation of B. subtilis) are structurally related and form a distinct class A of low-Mr PBPs/DD-peptidases. The distribution of the hydrophobic clusters along the amino acid sequences also shows that the Streptomyces K15 PBP, and by extension the other PBPs of class A, have similarity in the polypeptide folding, with the beta-lactamases of class A, with as reference the Streptomyces albus G and Staphylococcus aureus beta-lactamases of known three-dimensional structure. This comparison allows one to predict most of the secondary structures in the PBPs and the amino acid motifs that define the enzyme active sites.

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