U. Fristedt et al., Characterization of purified and unidirectionally reconstituted Pho84 phosphate permease of Saccharomyces cerevisiae, FEBS LETTER, 458(1), 1999, pp. 1-5
Hydropathy analysis of the amino acid sequence of the Pho84 phosphate perme
ase of Saccharomyces cerevisiae suggests that the protein consists of 12 tr
ansmembrane domains connected by hydrophilic loops. The Pho84 protein has b
een modified by a gene fusion approach, yielding two different N-terminal H
is-tagged chimeras which can be expressed in Escherichia coli, purified and
functionally reconstituted into defined proteoliposomes, The continuous ep
itopes in the N- and C-terminal sequences of the Pho84 chimeras were shown
to be accessible in proteoliposomes containing the purified active Pho84 pr
oteins. Site-specific proteolysis of the immunoreactive N-terminal sequence
in the reconstituted protein suggests a unidirectional insertion into lipo
somes. (C) 1999 Federation of European Biochemical Societies.