Staphylococcus aureus sortase A exists as a dimeric protein in vitro.
We report the first direct observation of the self-association behavior of the Staphylococcus aureus sortase A (SrtA) transpeptidase. Formation of a SrtA dimer was observed under native conditions by polyacrylamide gel electrophoresis and fast protein liquid chromatography (FPLC). Subsequent peptide mass fingerprinting and protein sequencing experiments confirmed the dimeric form of the SrtA protein. Furthermore, SrtA can be selectively cross-linked both in vitro and in Escherichia coli. Multiple samples of enzyme were subjected to analytical sedimentation equilibrium ultracentrifugation to obtain an apparent Kd for dimer formation of about 55 microM. Finally, enzyme kinetic studies suggested that the dimeric form of SrtA is more active than the monomeric enzyme. Discovery of SrtA dimerization may have significant implications for understanding microbial physiology and developing new antibiotics.
Lu
Changsheng
C
Division of Medicinal Chemistry, College of Pharmacy, University of Texas at Austin, Austin, Texas 78712, USA.
Zhu
Jie
J
Wang
Yun
Y
Umeda
Aiko
A
Cowmeadow
Roshani B
RB
Lai
Eric
E
Moreno
Gabrielle N
GN
Person
Maria D
MD
Zhang
Zhiwen
Z
D019881:Aminoacyltransferases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular; D003432:Cross-Linking Reagents / Q000737:chemistry; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D019281:Dimerization; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D007700:Kinetics; D010449:Peptide Mapping; D046928:Peptidoglycan Glycosyltransferase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D020539:Sequence Analysis, Protein; D013211:Staphylococcus aureus / Q000201:enzymology* / Q000235:genetics; D014461:Ultracentrifugation
D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D019881:Aminoacyltransferases; C120542:sortase A; D046928:Peptidoglycan Glycosyltransferase; D003546:Cysteine Endopeptidases