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The Life and Times of Lac Permease: Crystals Ain’t Everything, but They Certainly Do Help

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Membrane Transport Mechanism

Part of the book series: Springer Series in Biophysics ((BIOPHYSICS,volume 17))

Abstract

This chapter focuses on the lactose permease of Escherichia coli (LacY), a galactoside/H+ symporter, as this membrane transport protein is the grandparent for the major facilitator superfamily (MFS) and arguably the most intensively studied secondary transporter known at present.

LacY couples the free energy released from downhill translocation of H+ in response to an H+ electrochemical gradient to drive the stoichiometric accumulation of galactopyranosides against a concentration gradient under physiological conditions. X-ray structures of an inward-facing conformation and most recently, an almost occluded conformation with a narrow periplasmic opening have been solved, which confirm many conclusions from biochemical and biophysical studies. Although structure models are critical, they are not sufficient to explain the catalysis of transport. The clues to understanding transport mechanisms are based on the principles of enzyme kinetics. Secondary transport is a dynamic process that can be described only partially by the static snapshots provided by X-ray crystallography. However, without structural information it is virtually impossible to deduce the chemistry underlying ion-coupled transport. By combining a large body of biochemical/biophysical data derived from systematic studies of site-directed mutants in LacY, residues involved in substrate binding and H+ translocation have been identified. On the basis of the functional properties of the mutants and the X-ray structures, a working model for the symport mechanism that involves alternating access of the binding site is presented. The use of molecular biology to engineer LacY for dynamic studies combined with computational modeling has led to the postulate that the transport reaction is driven by thermodynamics, but is controlled kinetically.

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Acknowledgments

This work was supported by National Institutes of Health Grants DK51131, DK069463, and GM073210, as well as National Science Foundation Grant MCB-1129551 (to H.R.K.).

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Authors and Affiliations

  1. Department of Physiology, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA, 90095, USA

    M. Gregor Madej & H. Ronald Kaback

  2. Department of Microbiology, Immunology and Molecular Genetics, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA, 90095, USA

    H. Ronald Kaback

  3. Molecular Biology Institute, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA, 90095, USA

    H. Ronald Kaback

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  1. M. Gregor Madej
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  2. H. Ronald Kaback
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Correspondence to H. Ronald Kaback .

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  1. Department Chemistry Institute of Biochemistry, University of Cologne, Köln, Germany

    Reinhard Krämer

  2. Faculty for Biology and Preclinical Medicine, University of Regensburg, Regensburg, Germany

    Christine Ziegler

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Madej, M.G., Kaback, H.R. (2014). The Life and Times of Lac Permease: Crystals Ain’t Everything, but They Certainly Do Help. In: Krämer, R., Ziegler, C. (eds) Membrane Transport Mechanism. Springer Series in Biophysics, vol 17. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-53839-1_6

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