Purification of a Ran-interacting protein that is required for protein import into the nucleus.
Previously we reported the isolation of two cytosolic fractions (A and B) from Xenopus ovary that are required sequentially to support protein import into the nuclei of digitonin-permeabilized cells. Fraction A is required for recognition of the nuclear localization sequence and targeting to the nuclear envelope, whereas fraction B is required for the subsequent translocation of the bound substrate into the nucleus. The first protein required for fraction B activity to be purified was the small GTPase Ran (ras-related nuclear protein). Here we report the purification of the second (and final) protein required for fraction B activity. By SDS/PAGE, the purified protein appeared as a single band with an apparent molecular mass of 10 kDa, but the native protein fractionated upon gel filtration chromatography with an apparent size of 30 kDa. Peptide sequence analysis indicated that the purified protein was highly homologous to a previously identified human protein of unknown function called placental protein 15 (pp15) and to the predicted protein product of a yeast open reading frame from Saccharomyces cerevisiae.
Moore
M S
MS
Laboratory of Cell Biology, Howard Hughes Medical Institute, Rockefeller University, New York, NY 10021.
Blobel
G
G
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism*; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000378:metabolism; D005260:Female; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism*; D029543:Nucleocytoplasmic Transport Proteins; D010053:Ovary / Q000378:metabolism*; D051381:Rats; D011915:Rats, Inbred BUF; D029701:Saccharomyces cerevisiae Proteins; D029867:Xenopus Proteins; D014982:Xenopus laevis; D020931:ran GTP-Binding Protein
D002352:Carrier Proteins; C499531:NTF2 protein, S cerevisiae; D009687:Nuclear Proteins; D029543:Nucleocytoplasmic Transport Proteins; C089822:Ran-interacting protein p10, Xenopus; D029701:Saccharomyces cerevisiae Proteins; D029867:Xenopus Proteins; D019204:GTP-Binding Proteins; D020931:ran GTP-Binding Protein