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Crystallization and preliminary crystallographic analysis of the transpeptidase domain of penicillin-binding protein 2B from Streptococcus pneumoniae.

ABSTRACT: Penicillin-binding protein (PBP) 2B from Streptococcus pneumoniae catalyzes the cross-linking of peptidoglycan precursors that occurs during bacterial cell-wall biosynthesis. A selenomethionyl (SeMet) substituted PBP 2B transpeptidase domain was isolated from a limited proteolysis digest of a soluble form of recombinant PBP 2B and then crystallized. The crystals belonged to space group P4(3)2(1)2, with unit-cell parameters a = b = 86.39, c = 143.27 A. Diffraction data were collected to 2.4 A resolution using the BL32B2 beamline at SPring-8. The asymmetric unit contains one protein molecule and 63.7% solvent.

SUBMITTER: Yamada M

PROVIDER: S-EPMC2374250 | biostudies-literature | 2008 Apr

REPOSITORIES: biostudies-literature

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Crystallization and preliminary crystallographic analysis of the transpeptidase domain of penicillin-binding protein 2B from Streptococcus pneumoniae.

Yamada Mototsugu M Watanabe Takashi T Baba Nobuyoshi N Miyara Takako T Saito Jun J Takeuchi Yasuo Y

Acta crystallographica. Section F, Structural biology and crystallization communications 20080321 Pt 4

Penicillin-binding protein (PBP) 2B from Streptococcus pneumoniae catalyzes the cross-linking of peptidoglycan precursors that occurs during bacterial cell-wall biosynthesis. A selenomethionyl (SeMet) substituted PBP 2B transpeptidase domain was isolated from a limited proteolysis digest of a soluble form of recombinant PBP 2B and then crystallized. The crystals belonged to space group P4(3)2(1)2, with unit-cell parameters a = b = 86.39, c = 143.27 A. Diffraction data were collected to 2.4 A res ...[more]

PMID: 18391428

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Project description:Unlabelled?-Lactam antibiotics are the drugs of choice to treat pneumococcal infections. The spread of ?-lactam-resistant pneumococci is a major concern in choosing an effective therapy for patients. Systematically tracking ?-lactam resistance could benefit disease surveillance. Here we developed a classification system in which a pneumococcal isolate is assigned to a "PBP type" based on sequence signatures in the transpeptidase domains (TPDs) of the three critical penicillin-binding proteins (PBPs), PBP1a, PBP2b, and PBP2x. We identified 307 unique PBP types from 2,528 invasive pneumococcal isolates, which had known MICs to six ?-lactams based on broth microdilution. We found that increased ?-lactam MICs strongly correlated with PBP types containing divergent TPD sequences. The PBP type explained 94 to 99% of variation in MICs both before and after accounting for genomic backgrounds defined by multilocus sequence typing, indicating that genomic backgrounds made little independent contribution to ?-lactam MICs at the population level. We further developed and evaluated predictive models of MICs based on PBP type. Compared to microdilution MICs, MICs predicted by PBP type showed essential agreement (MICs agree within 1 dilution) of >98%, category agreement (interpretive results agree) of >94%, a major discrepancy (sensitive isolate predicted as resistant) rate of <3%, and a very major discrepancy (resistant isolate predicted as sensitive) rate of <2% for all six ?-lactams. Thus, the PBP transpeptidase signatures are robust indicators of MICs to different ?-lactam antibiotics in clinical pneumococcal isolates and serve as an accurate alternative to phenotypic susceptibility testing.ImportanceThe human pathogen Streptococcus pneumoniae is a leading cause of morbidity and mortality worldwide. ?-Lactam antibiotics such as penicillin and ceftriaxone are the drugs of choice to treat pneumococcal infections. Some pneumococcal strains have developed ?-lactam resistance through altering their penicillin-binding proteins (PBPs) and have become a major concern in choosing effective patient therapy. To systematically track and predict ?-lactam resistance, we obtained the sequence signatures of PBPs from a large collection of clinical pneumococcal isolates using whole-genome sequencing data and found that these "PBP types" were predictive of resistance levels. Our findings can benefit the current era of strain surveillance when whole-genome sequencing data often lacks detailed resistance information. Using PBP positions that we found are always substituted within highly resistant strains may lead to further refinements. Sequence-based predictions are accurate and may lead to the ability to extract critical resistance information from nonculturable clinical specimens.

Dataset Information

Crystallization and preliminary crystallographic analysis of the transpeptidase domain of penicillin-binding protein 2B from Streptococcus pneumoniae.

Publications

Crystallization and preliminary crystallographic analysis of the transpeptidase domain of penicillin-binding protein 2B from Streptococcus pneumoniae.

Similar Datasets

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