Gabrielle N Moreno

Publications (2)6.84 Total impact

• Article: Single mutation on the surface of Staphylococcus aureus Sortase A can disrupt its dimerization.

  Jie Zhu, Changsheng Lu, Matthew Standland, Eric Lai, Gabrielle N Moreno, Aiko Umeda, Xudong Jia, Zhiwen Zhang
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  ABSTRACT: Staphylococcus aureus Sortase A (SrtA) is an important Gram-positive membrane enzyme which catalyzes the anchoring of many cell surface proteins conserved with the LPXTG sequence. Recently SrtA has been demonstrated to be a dimer with a Kd of 55 microM in vitro. Herein, we show that a single point mutation of amino acid residue on the surface of SrtA can completely disrupt the dimerization. Native polyacrylamide gel electrophoresis and analytical gel filtration chromatography were used to detect the dimer-monomer equilibrium of SrtA mutants. Circular dichroism spectrum experiments were performed to study the conformational change of each SrtA mutant. An enzyme activity assay confirmed that all the SrtA mutants were active in vitro. Our results not only are important for understanding the SrtA protein self-associating mechanism but also provided the necessary starting materials for the study of sortase A pathway in vivo, which may have significant implications for discovering microbial physiology and give a potential target for novel Gram-positive antibiotics.
  Biochemistry 03/2008; 47(6):1667-74. · 3.42 Impact Factor
• Article: Staphylococcus aureus sortase A exists as a dimeric protein in vitro.

  Changsheng Lu, Jie Zhu, Yun Wang, Aiko Umeda, Roshani B Cowmeadow, Eric Lai, Gabrielle N Moreno, Maria D Person, Zhiwen Zhang
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  ABSTRACT: 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.
  Biochemistry 09/2007; 46(32):9346-54. · 3.42 Impact Factor