STRUCTURAL TOPOLOGY OF TRANSMEMBRANE HELIX-10 IN THE LACTOSE PERMEASEOF ESCHERICHIA-COLI

In the lactose permease of Escherichia coli, transmembrane helix 10 ha s been shown to be functionally important, The structure of this helix has been examined in greater detail in this study, A total of 46 subs titution and 8 insertional mutants were constructed and analyzed along the entire length of transmembrane helix 10, The results identified a mino acids that are tolerant of substitutions by a variety of amino ac ids, Since a number of these amino acids (Thr-320, Val-331, Phe-325, a nd Ile-317) are clustered in one region in a helical wheel projection of transmembrane helix 10, it seems likely that this face of helix 10 is interacting with the membrane, The channel lining domain is thought to consist of the helical face containing Glu-325, Leu-318, Leu-329, His-322, Val-315, Cys-333, Val-326, and Lys-319 based on the results h ere and from earlier findings, Deleterious mutations along this face t ended to greatly increase the K-m value for lactose transport with onl y minor effects on the V-max. Analysis of insertional mutants revealed that perturbation of the spatial relationship between amino acids at the periplasmic edge is less deleterious than perturbation in the cent er of the helix or the cytoplasmic edge. Using all of the above inform ation, a detailed structural topology of transmembrane helix 10 is pro posed.