Structural characterization of Saccharomyces cerevisiae prion-like protein Ure2

被引:109
作者
Thual, C
Komar, AA
Bousset, L
Fernandez-Bellot, E
Cullin, C
Melki, R [1 ]
机构
[1] CNRS, Lab Enzymol & Biochim Struct, F-91198 Gif Sur Yvette, France
[2] CNRS, Ctr Genet Mol, F-91198 Gif Sur Yvette, France
关键词
D O I
10.1074/jbc.274.19.13666
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Sacchromyces cerevisiae prion-like protein Urea was expressed in Escherichia coli and was purified to homogeneity, We show here that Ure2p is a soluble protein that can assemble into fibers that are similar to the fibers observed in the case of PrP in its scrapie prion filaments form or that form on Sup35 self-assembly. Ure2p self-assembly is a cooperative process where one can distinguish a lag phase followed by an elongation phase preceding a plateau. A combination of size exclusion chromatography, sedimentation velocity, and electron microscopy demonstrates that the soluble form of Ure2p consists at least of three forms of the protein as follows: a monomeric, dimeric, and tetrameric form whose abundance is concentration-dependent. By the use of limited proteolysis, intrinsic fluorescence, and circular dichroism measurements, we bring strong evidence for the existence of at least two structural domains in Ure2p molecules. Indeed, Ure2p NH2-terminal region is found poorly structured, whereas its COOH-terminal domain appears to be compactly folded. Finally, we show that only slight conformational changes accompany Ure2p assembly into insoluble high molecular weight oligomers, These changes essentially affect the COOH-terminal part of the molecule. The properties of Ure2p are compared in the discussion to that of other prion-like proteins such as Sup35 and mammalian prion protein PrP.
引用
收藏
页码:13666 / 13674
页数:9
相关论文
共 41 条
[1]  
BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3
[2]   Pathologic conformations of prion proteins [J].
Cohen, FE ;
Prusiner, SB .
ANNUAL REVIEW OF BIOCHEMISTRY, 1998, 67 :793-+
[3]   THE URE2 GENE-PRODUCT OF SACCHAROMYCES-CEREVISIAE PLAYS AN IMPORTANT ROLE IN THE CELLULAR-RESPONSE TO THE NITROGEN-SOURCE AND HAS HOMOLOGY TO GLUTATHIONE S-TRANSFERASES [J].
COSCHIGANO, PW ;
MAGASANIK, B .
MOLECULAR AND CELLULAR BIOLOGY, 1991, 11 (02) :822-832
[4]   Structure and functions of the 20S and 26S proteasomes [J].
Coux, O ;
Tanaka, K ;
Goldberg, AL .
ANNUAL REVIEW OF BIOCHEMISTRY, 1996, 65 :801-847
[5]   IDENTIFICATION OF PRION AMYLOID FILAMENTS IN SCRAPIE-INFECTED BRAIN [J].
DEARMOND, SJ ;
MCKINLEY, MP ;
BARRY, RA ;
BRAUNFELD, MB ;
MCCOLLOCH, JR ;
PRUSINER, SB .
CELL, 1985, 41 (01) :221-235
[6]   Structure of the recombinant full-length hamster prion protein PrP(29-231): The N terminus is highly flexible [J].
Donne, DG ;
Viles, JH ;
Groth, D ;
Mehlhorn, I ;
James, TL ;
Cohen, FE ;
Prusiner, SB ;
Wright, PE ;
Dyson, HJ .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1997, 94 (25) :13452-13457
[7]   Characterization of the interaction domains of Ure2p, a prion-like protein of yeast [J].
Fernandez-Bellot, E ;
Guillemet, E ;
Baudin-Baillieu, A ;
Gaumer, S ;
Komar, AA ;
Cullin, C .
BIOCHEMICAL JOURNAL, 1999, 338 :403-407
[8]   Self-seeded fibers formed by Sup35, the protein determinant of [PSI+], a heritable prion-like factor of S-cerevisiae [J].
Glover, JR ;
Kowal, AS ;
Schirmer, EC ;
Patino, MM ;
Liu, JJ ;
Lindquist, S .
CELL, 1997, 89 (05) :811-819
[9]   Deadly conformations - Protein misfolding in prion disease [J].
Horwich, AL ;
Weissman, JS .
CELL, 1997, 89 (04) :499-510
[10]   CHARACTERIZATION OF MESSENGER-RNA FOR A PROLINE-RICH PROTEIN OF COTTON FIBER [J].
JOHN, ME ;
KELLER, G .
PLANT PHYSIOLOGY, 1995, 108 (02) :669-676