In:
Proceedings of the National Academy of Sciences, Proceedings of the National Academy of Sciences, Vol. 114, No. 7 ( 2017-02-14), p. 1530-1535
Abstract:
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 (pK app ) 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 pK a 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.
Type of Medium:
Online Resource
ISSN:
0027-8424
,
1091-6490
DOI:
10.1073/pnas.1621431114
Language:
English
Publisher:
Proceedings of the National Academy of Sciences
Publication Date:
2017
detail.hit.zdb_id:
209104-5
detail.hit.zdb_id:
1461794-8
SSG:
11
SSG:
12
