© 2001 Macmillan Magazines Ltd 610 | AUGUST 2001 | VOLUME 2 www.nature.com/reviews/molcellbio REVIEWS Of the genomes sequenced so far, over 20% of the gene products are known or predicted to be polytopic trans- membrane proteins 1 . These proteins catalyse a multi- tude of essential functions, one of which is the transport of molecules into and out of cells or intracellular organelles, and across epithelia. However, membrane proteins in general — and membrane transport pro- teins in particular — are notoriously resistant to the determination of high-resolution structure by tradi- tional means because of their hydrophobicity and, in many instances, because of their metastable nature. Although advances in molecular biology and bio- chemistry have led to rapid progress in understanding structure–function relationships for some membrane proteins, structures have been obtained at atomic reso- lution in only a handful of instances. So, the level of understanding of membrane proteins is almost inverse- ly proportional to their roles in living systems. As more genomes are sequenced and a growing number of membrane proteins are identified, this discrepancy will probably increase. Moreover, in the post-genomic era, with proteomics emerging as a new field, limitations in studying large, hydrophobic membrane proteins will become even more acute. Transport proteins are a principal class of integral membrane proteins, many of which transduce the free energy stored in ELECTROCHEMICAL H + GRADIENTS into sub- strate concentration gradients across a membrane. By contrast,channel proteins — another important class of membrane proteins — do not transduce energy, but function as selective pores that often open in response to a specific stimulus to move solute down an electro- chemical H + gradient 2 . Like channels, membrane-trans- port proteins are highly relevant to human physiology and disease (BOX 1) . Furthermore, at least two of the most widely prescribed drugs in the world, fluoxetine (Prozac) and omeprazole (Prilosec), are targeted to membrane-transport proteins. The lactose permease One large group of evolutionarily related transport pro- teins is the major facilitator superfamily (MFS; BOX 2) 3 . Members of this family are found in membranes from archaea to the mammalian central nervous system, and they catalyse the transport of various solutes. An impor- tant model for the MFS, as well as for other membrane proteins, is the lactose permease in the Escherichia coli cytoplasmic membrane, which is encoded by lacY,the second structural gene of the lac operon 4 . Primary and secondary structure. LacY was the first gene encoding a membrane-transport protein to be cloned and sequenced 5 . This led to the overexpression 6 , solubilization 7 and purification of lactose permease in a completely active state (reviewed in REF. 8), as well as the demonstration that the protein functions as a monomer (see REF. 9). So, all properties of the β-galacto- side transport system in E. coli can be attributed to a single gene product. The lactose permease is composed of 417 amino- acid residues, is ~70% helical 10,11 , and has 12 helices that traverse the membrane in zig–zag fashion connected by THE KAMIKAZE APPROACH TO MEMBRANE TRANSPORT H. Ronald Kaback, Miklós Sahin-Tóth and Adam B. Weinglass Membrane transport proteins catalyse the movement of molecules into and out of cells and organelles, but their hydrophobic and metastable nature often makes them difficult to study by traditional means. Novel approaches that have been developed and applied to one membrane transport protein, the lactose permease from Escherichia coli , are now being used to study various other membrane proteins. Howard Hughes Medical Institute, Departments of Physiology, and Microbiology, Immunology and Molecular Genetics, and the Molecular Biology Institute, University of California, Los Angeles, California 90095-1662, USA. Correspondence to H.R.K. e-mail: ronaldk@hhmi.ucla.edu ELECTROCHEMICAL H + GRADIENT (∆μ H+ ).When two aqueous phases are separated by a membrane, the electrochemical potential difference of H + between the two phases is expressed as ∆μ H+ /F = ∆Ψ - 2.3RT/F∆pH, where F is the Faraday constant, ∆Ψ is the electric potential difference between two phases, R is the gas constant, T is the absolute temperature,and ∆pH is the difference in the concentration of H + across the membrane.