Structural topology of transmembrane helix 10 in the lactose permease of Escherichia coli.
In the lactose permease of Escherichia coli, transmembrane helix 10 has been shown to be functionally important. The structure of this helix has been examined in greater detail in this study. A total of 46 substitution and 8 insertional mutants were constructed and analyzed along the entire length of transmembrane helix 10. The results identified amino acids that are tolerant of substitutions by a variety of amino acids. Since a number of these amino acids (Thr-320, Val-331, Phe-325, and 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 here and from earlier findings. Deleterious mutations along this face tended to greatly increase the Km value for lactose transport with only minor effects on the Vmax. 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 center of the helix or the cytoplasmic edge. Using all of the above information, a detailed structural topology of transmembrane helix 10 is proposed.
Goswitz
V C
VC
Department of Genetics and Cell Biology, University of Minnesota, St. Paul, Minnesota 55108, USA.
Matzke
E A
EA
Taylor
M R
MR
Jessen-Marshall
A E
AE
Brooker
R J
RJ
D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D005697:Galactosides / Q000378:metabolism; D018257:Helix-Loop-Helix Motifs; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D027981:Symporters
D029968:Escherichia coli Proteins; D005697:Galactosides; C050221:LacY protein, E coli; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D027981:Symporters; C021499:lactose permease