Analysis of the pore of the unusual major intrinsic protein channel, yeast Fps1p.
Journal article, 2001
Fps1p is a glycerol efflux channel from Saccharomyces cerevisiae. In this atypical major intrinsic protein neither of the signature NPA motifs of the family, which are part of the pore, is preserved. To understand the functional consequences of this feature, we analyzed the pseudo-NPA motifs of Fps1p by site-directed mutagenesis and assayed the resultant mutant proteins in vivo. In addition, we took advantage of the fact that the closest bacterial homolog of Fps1p, Escherichia coli GlpF, can be functionally expressed in yeast, thus enabling the analysis in yeast cells of mutations that make this typical major intrinsic protein more similar to Fps1p. We observed that mutations made in Fps1p to "restore" the signature NPA motifs did not substantially affect channel function. In contrast, when GlpF was mutated to resemble Fps1p, all mutants had reduced activity compared with wild type. We rationalized these data by constructing models of one GlpF mutant and of the transmembrane core of Fps1p. Our model predicts that the pore of Fps1p is more flexible than that of GlpF. We discuss the fact that this may accommodate the divergent NPA motifs of Fps1p and that the different pore structures of Fps1p and GlpF may reflect the physiological roles of the two glycerol facilitators.
Escherichia coli
metabolism
metabolism
Time Factors
Amino Acid Motifs
Immunoblotting
Membrane Proteins
Protein Structure
metabolism
Escherichia coli Proteins
Saccharomyces cerevisiae
Glycerol
Point Mutation
Mutation
Tertiary
chemistry
Saccharomyces cerevisiae Proteins
DNA Primers
metabolism
Mutagenesis
Aquaporins
Fungal Proteins
Plasmids
Cell Membrane
chemistry
chemistry
Molecular
Site-Directed
metabolism
Models
Bacterial Outer Membrane Proteins
metabolism
Protein Binding
Author
R Bill
University of Gothenburg
Kristina Hedfalk
Chalmers, Department of Chemistry and Bioscience
S Karlgren
J G Mullins
J Rydström
S Hohmann
Journal of Biological Chemistry
0021-9258 (ISSN) 1083-351X (eISSN)
Vol. 276 39 36543-9Subject Categories (SSIF 2011)
Biological Sciences
DOI
10.1074/jbc.M105045200
PubMed
11445582