title: Properties of a novel PBP2A protein homolog from Staphylococcus aureus strain LGA251 and its contribution to the β-lactam-resistant phenotype.
abstract: Methicillin-resistant Staphylococcus aureus (MRSA) strains show strain-to-strain variation in resistance level, in genetic background, and also in the structure of the chromosomal cassette (SCCmec) that carries the resistance gene mecA. In contrast, strain-to-strain variation in the sequence of the mecA determinant was found to be much more limited among MRSA isolates examined so far. The first exception to this came with the recent identification of MRSA strain LGA251, which carries a new homolog of this gene together with regulatory elements mecI/mecR that also have novel, highly divergent structures. After cloning and purification in Escherichia coli, PBP2A(LGA), the protein product of the new mecA homolog, showed aberrant mobility in SDS-PAGE, structural instability and loss of activity at 37 °C, and a higher relative affinity for oxacillin as compared with cefoxitin. The mecA homolog free of its regulatory elements was cloned into a plasmid and introduced into the background of the β-lactam-susceptible S. aureus strain COL-S. In this background, the mecA homolog expressed a high-level resistance to cefoxitin (MIC = 400 μg/ml) and a somewhat lower resistance to oxacillin (minimal inhibitory concentration = 200 μg/ml). Similar to PBP2A, the protein homolog PBP2A(LGA) was able to replace the essential function of the S. aureus PBP2 for growth. In contrast to PBP2A, PBP2A(LGA) did not depend on the transglycosylase activity of the native PBP2 for expression of high level resistance to oxacillin, suggesting that the PBP2A homolog may preferentially cooperate with a monofunctional transglycosylase as the alternative source of transglycosylase activity.
authors.affiliation: Laboratory of Microbiology and Infectious Diseases, The Rockefeller University, New York, New York 10065, USA.
authors.forename: Choonkeun
authors.initials: C
authors.lastname: Kim
authors.forename: Catarina
authors.initials: C
authors.lastname: Milheiriço
authors.forename: Susana
authors.initials: S
authors.lastname: Gardete
authors.forename: Mark A
authors.initials: MA
authors.lastname: Holmes
authors.forename: Matt T G
authors.initials: MT
authors.lastname: Holden
authors.forename: Hermínia
authors.initials: H
authors.lastname: de Lencastre
authors.forename: Alexander
authors.initials: A
authors.lastname: Tomasz
mesh_terms: D019881:Aminoacyltransferases / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D001896:Boron Compounds / Q000737:chemistry; D002440:Cefoxitin / Q000737:chemistry / Q000494:pharmacology; D002462:Cell Membrane / Q000737:chemistry; D003001:Cloning, Molecular; D004795:Enzyme Stability; D020128:Inhibitory Concentration 50; D008826:Microbial Sensitivity Tests; D008970:Molecular Weight; D010068:Oxacillin / Q000737:chemistry / Q000494:pharmacology; D046915:Penicillin-Binding Proteins; D010406:Penicillins / Q000737:chemistry; D010641:Phenotype; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D013211:Staphylococcus aureus / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D018440:beta-Lactam Resistance / Q000235:genetics*
chemical_list: C118961:BOCILLIN FL; D001426:Bacterial Proteins; D001896:Boron Compounds; D046915:Penicillin-Binding Proteins; D010406:Penicillins; C487789:mecA protein, Staphylococcus aureus; D002440:Cefoxitin; D019881:Aminoacyltransferases; C022330:peptidoglycan transpeptidase; D010068:Oxacillin