Article

Structural determination of wild-type lactose permease.

Department of Physiology, University of California, Los Angeles, CA 90095-1662, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.81). 10/2007; 104(39):15294-8. DOI: 10.1073/pnas.0707688104
Source: PubMed

ABSTRACT 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 six-helix bundles with twofold pseudosymmetry separated by a large interior hydrophilic cavity open only to the cytoplasmic side and containing the side chains important for sugar and H(+) binding. To initiate transport, binding of sugar and/or an H(+) electrochemical gradient increases the probability of opening on the periplasmic side. Because the inward-facing conformation represents the lowest free-energy state, the rate-limiting step for transport may be the conformational change leading to the outward-facing conformation.

0 Bookmarks
 · 
151 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: Soluble sugars serve five main purposes in multicellular organisms: as sources of carbon skeletons, osmolytes, signals, and transient energy storage and as transport molecules. Most sugars are derived from photosynthetic organisms, particularly plants. In multicellular organisms, some cells specialize in providing sugars to other cells (e.g., intestinal and liver cells in animals, photosynthetic cells in plants), whereas others depend completely on an external supply (e.g., brain cells, roots and seeds). This cellular exchange of sugars requires transport proteins to mediate uptake or release from cells or subcellular compartments. Thus, not surprisingly, sugar transport is critical for plants, animals, and humans. At present, three classes of eukaryotic sugar transporters have been characterized, namely the glucose transporters (GLUTs), sodium-glucose symporters (SGLTs), and SWEETs. This review presents the history and state of the art of sugar transporter research, covering genetics, biochemistry, and physiology-from their identification and characterization to their structure, function, and physiology. In humans, understanding sugar transport has therapeutic importance (e.g., addressing diabetes or limiting access of cancer cells to sugars), and in plants, these transporters are critical for crop yield and pathogen susceptibility. Expected final online publication date for the Annual Review of Biochemistry Volume 84 is June 02, 2015. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: In a variety of bacteria, the phosphotransferase protein IIAGlc plays a key regulatory role in catabolite repression in addition to its role in the vectorial phosphorylation of glucose catalyzed by the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS). The lactose permease (LacY) of Escherichia coli catalyzes stoichiometric symport of a galactoside with an H+, using a mechanism in which sugar- and H+-binding sites become alternatively accessible to either side of the membrane. Both the expression (via regulation of cAMP levels) and the activity of LacY are subject to regulation by IIAGlc (inducer exclusion). Here we report the thermodynamic features of the IIAGlc-LacY interaction as measured by isothermal titration calorimetry (ITC). The studies show that IIAGlc binds to LacY with a Kd of about 5 muM and a stoichiometry of unity and that binding is driven by solvation entropy and opposed by enthalpy. Upon IIAGlc binding, the conformational entropy of LacY is restrained, which leads to a significant decrease in sugar affinity. By suppressing conformational dynamics, IIAGlc blocks inducer entry into cells and favors constitutive glucose uptake and utilization. Furthermore, the studies support the notion that sugar binding involves an induced-fit mechanism that is inhibited by IIAGlc binding. The precise mechanism of the inhibition of LacY by IIAGlc elucidated by ITC differs from the inhibition of melibiose permease (MelB), supporting the idea that permeases can differ in their thermodynamic response to binding IIAGlc.
    Proceedings of the National Academy of Sciences 02/2015; DOI:10.1073/pnas.1500891112 · 9.81 Impact Factor
  • Source

Full-text (2 Sources)

Download
37 Downloads
Available from
Jun 4, 2014