pK a of Glu325 in LacY

Autor: Petra Hellwig, H. Ronald Kaback, Junichi Sugihara, Natalia Grytsyk
Přispěvatelé: Laboratoire de Bioimagerie et Pathologies (LBP), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS), Biophotonique et Pharmacologie (CNRS UMR 7213), Centre National de la Recherche Scientifique (CNRS), Chimie de la matière complexe (CMC), Université de Strasbourg (UNISTRA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)
Rok vydání: 2017
Předmět:
Zdroj: Proceedings of the National Academy of Sciences of the United States of America
Proceedings of the National Academy of Sciences of the United States of America, National Academy of Sciences, 2017, 114 (7), pp.1530-1535. ⟨10.1073/pnas.1621431114⟩
ISSN: 1091-6490
0027-8424
Popis: Lactose permease (LacY), a paradigm for the largest family of membrane transport proteins, catalyzes the coupled translocation of a galactoside and a H+ across the cytoplasmic membrane of Escherichia coli (galactoside/H+ symport). One of the most important aspects of the mechanism is the relationship between protonation and binding of the cargo galactopyranoside. In this regard, it has been shown that protonation is required for binding. Furthermore when galactoside affinity is measured as a function of pH, an apparent pK (pKapp) of ∼10.5 is obtained. Strikingly, when Glu325, a residue long known to be involved in coupling between H+ and sugar translocation, is replaced with a neutral side chain, the pH effect is abolished, and high-affinity binding is observed until LacY is destabilized at alkaline pH. In this paper, infrared spectroscopy is used to identify Glu325 in situ. Moreover, it is demonstrated that this residue exhibits a pKa of 10.5 ± 0.1 that is insensitive to the presence of galactopyranoside. Thus, it is apparent that protonation of Glu325 specifically is required for effective sugar binding to LacY.
Databáze: OpenAIRE
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