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An X-ray structure of the lactose permease of Escherichia coli (LacY) in an inward-facing conformation has been solved. LacY contains N- and C-terminal domains, each with six transmembrane helices, positioned pseudosymmetrically. Ligand is bound at the apex of a hydrophilic cavity in the approximate middle of the molecule. Residues involved in substrate(More)
The understanding of integral membrane protein (IMP) structure and function is hampered by the difficulty of handling these proteins. Aqueous solubilization, necessary for many types of biophysical analysis, generally requires a detergent to shield the large lipophilic surfaces of native IMPs. Many proteins remain difficult to study owing to a lack of(More)
Here we describe an x-ray structure of wild-type lactose permease (LacY) from Escherichia coli determined by manipulating phospholipid content during crystallization. The structure exhibits the same global fold as the previous x-ray structures of a mutant that binds sugar but cannot catalyze translocation across the membrane. LacY is organized into two(More)
Cation-coupled active transport is an essential cellular process found ubiquitously in all living organisms. Here, we present two novel ligand-free X-ray structures of the lactose permease (LacY) of Escherichia coli determined at acidic and neutral pH, and propose a model for the mechanism of coupling between lactose and H+ translocation. No sugar-binding(More)
The bacterial melibiose permease (MelB) belongs to the glycoside-pentoside-hexuronide:cation symporter family, a part of the major facilitator superfamily (MFS). Structural information regarding glycoside-pentoside-hexuronide:cation symporter family transporters and other Na(+)-coupled permeases within MFS has been lacking, although a wealth of biochemical(More)
This protocol describes a detailed method to study the static and dynamic features of membrane proteins, as well as solvent accessibility, by utilizing the lactose permease of Escherichia coli (LacY) as a model. The method relies on the use of functional single-Cys mutants, an affinity tag and a PhosphoImager. The membrane-permeant, radioactive thiol(More)
The melibiose permease of Escherichia coli (MelB) catalyzes the coupled stoichiometric symport of a galactoside with a cation (either Na(+), Li(+), or H(+)), using free energy from the downhill translocation of one cosubstrate to catalyze the accumulation of the other. Here, we present a 3D structure model of MelB threaded through a crystal structure of the(More)
BACKGROUND/AIMS In the present study, we explored reactive axygen species (ROS) production in mitochondria, the mechanism of hexavalent chromium (Cr(VI)) hepatotoxicity, and the role of protection by GSH. METHODS Intact mitochondria were isolated from rat liver tissues and mitochondrial basal respiratory rates of NADH and FADH2 respiratory chains were(More)
The application of immunoprotein-based targeting strategies to the boron neutron-capture therapy of cancer poses an exceptional challenge, because viable boron neutron-capture therapy by this method will require the efficient delivery of 10(3) boron-10 atoms by each antigen-binding protein. Our recent investigations in this area have been focused on the(More)
Crystallization is a major bottleneck to obtaining x-ray structures of membrane proteins. By applying an established crystallization protocol for the lactose permease (LacY) of Escherichia coli, a systematic study of the effect of phospholipids (PL) on crystallization of LacY was undertaken. We observe three different crystal forms that diffract to(More)