Identification, purification, and characterization of transpeptidase and glycosyltransferase domains of Streptococcus pneumoniae penicillin-binding protein 1a

被引:46
作者
Di Guilmi, AM
Mouz, N
Andrieu, JP
Hoskins, J
Jaskunas, SR
Gagnon, J
Dideberg, O
Vernet, T
机构
[1] Inst Biol Struct Jean Pierre Ebel, Lab Ingn Macromolecules, CEA, CNRS, F-38027 Grenoble 1, France
[2] Eli Lilly & Co, Lilly Res Labs, Indianapolis, IN 46285 USA
关键词
D O I
10.1128/JB.180.21.5652-5659.1998
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
Resistance to beta-lactam antibiotics in Streptococcus pneumoniae is due to alteration of penicillin-binding proteins (PBPs). S. pneumoniae PBP 1a belongs to the class A high-molecular-mass PBPs, which harbor transpeptidase (TP) and glycosyltransferase (GT) activities. The GT active site represents a new potential target for the generation of novel nonpenicillin antibiotics, The 683-amino-acid extracellular region of PBP 1a (PBP 1a*) was expressed in Escherichia coli as a GST fusion protein. The GST-PBP 1a* soluble protein was purified, and its domain organization was revealed by limited proteolysis, A protease-resistant fragment spanning Ser 264 to Arg 653 exhibited a reactivity profile against both beta-lactams and substrate analogues similar to that of the parent protein, This protein fragment represents the TP domain. The GT domain (Ser 37 to Lys 263) was expressed as a recombinant GST fusion protein, Protection by moenomycin of the GT domain against trypsin degradation was interpreted as an interaction between the GT domain and the moenomycin.
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页码:5652 / 5659
页数:8
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