Location of subunit-subunit contact sites on RNA polymerase II subunit 3 from the fission yeast Schizosaccharomyces pombe

被引:14
作者
Yasui, K
Ishiguro, A
Ishihama, A [1 ]
机构
[1] Natl Inst Genet, Dept Mol Genet, Shizuoka 4118540, Japan
[2] Grad Univ Adv Studies, Sch Life Sci, Shizuoka 4118540, Japan
关键词
D O I
10.1021/bi972939b
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
RNA polymerase II from the fission yeast Schizosaccharomyces pombe consists of 10 putative subunits. Subunit 3 (Rpb3) is a homologue of prokaryotic alpha subunit, which plays a key role in the assembly of core enzyme subunits. Previously we indicated that Rpb3 also plays an essential role in subunit assembly because it interacts with at least four subunits, two large subunits (Rpb1 and Rpb2) and two medium-sized subunits (Rpb3 and Rpb5) (1), and it constitutes a core subassembly consisting of Rpb2, Rpb3, and Rpb11 (2). Using a synthetic mixture of equimolar amounts of individual subunits, which were all purified from cDNA-expressed Eschelichia coli, we found here that Rpb3 also interacts with Rpb11, another alpha homologue. By making a set of Rpb3 deletion derivatives, we carried out mapping of the Rpb5- and Rpb11-contact sites on Rpb3. By far-Western blot and GST pull-down assays, we found that the amino acid sequence between residues 105-263 of Rpb3 is involved in binding Rpb5, and the sequence between residues 105-297 is required for binding Rpb11, Although the Rpb5- and Rpb11-contact sites on Rpb3 overlap each other, both subunits are able to associate with Rpb3 simultaneously. The binding of Rpb5 stabilizes the Rpb3-Rpb11 heterodimer.
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页码:5542 / 5548
页数:7
相关论文
共 28 条
[1]   SUBUNITS OF THE SCHIZOSACCHAROMYCES-POMBE RNA POLYMERASE-II - ENZYME-PURIFICATION AND STRUCTURE OF THE SUBUNIT 3 GENE [J].
AZUMA, Y ;
YAMAGISHI, M ;
ISHIHAMA, A .
NUCLEIC ACIDS RESEARCH, 1993, 21 (16) :3749-3754
[2]   CLONING AND SEQUENCE DETERMINATION OF THE SCHIZOSACCHAROMYCES-POMBE RPB1 GENE ENCODING THE LARGEST SUBUNIT OF RNA POLYMERASE-II [J].
AZUMA, Y ;
YAMAGISHI, M ;
UESHIMA, R ;
ISHIHAMA, A .
NUCLEIC ACIDS RESEARCH, 1991, 19 (03) :461-468
[3]   BIPARTITE FUNCTIONAL MAP OF THE ESCHERICHIA-COLI RNA POLYMERASE-ALPHA SUBUNIT - INVOLVEMENT OF THE C-TERMINAL REGION IN TRANSCRIPTION ACTIVATION BY CAMP-CRP [J].
IGARASHI, K ;
ISHIHAMA, A .
CELL, 1991, 65 (06) :1015-1022
[4]  
Ishihama A, 1981, Adv Biophys, V14, P1
[5]  
Ishihama A., 1997, NUCL ACIDS MOL BIOL, V11, P53
[6]   CLONING AND SEQUENCE DETERMINATION OF THE SCHIZOSACCHAROMYCES-POMBE RPB2 GENE ENCODING THE SUBUNIT-2 OF RNA POLYMERASE-II [J].
KAWAGISHI, M ;
YAMAGISHI, M ;
ISHIHAMA, A .
NUCLEIC ACIDS RESEARCH, 1993, 21 (03) :469-473
[7]   RNA polymerase II subunits 2, 3, and 11 form a core subassembly with DNA binding activity [J].
Kimura, M ;
Ishiguro, A ;
Ishihama, A .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1997, 272 (41) :25851-25855
[8]   FUNCTIONAL MAP OF THE ALPHA-SUBUNIT OF ESCHERICHIA-COLI RNA-POLYMERASE - INSERTION ANALYSIS OF THE AMINO-TERMINAL ASSEMBLY DOMAIN [J].
KIMURA, M ;
ISHIHAMA, A .
JOURNAL OF MOLECULAR BIOLOGY, 1995, 248 (04) :756-767
[9]   FUNCTIONAL MAP OF THE ALPHA-SUBUNIT OF ESCHERICHIA-COLI RNA-POLYMERASE - DELETION ANALYSIS OF THE AMINO-TERMINAL ASSEMBLY DOMAIN [J].
KIMURA, M ;
FUJITA, N ;
ISHIHAMA, A .
JOURNAL OF MOLECULAR BIOLOGY, 1994, 242 (02) :107-115
[10]   FUNCTIONAL MAP OF THE ALPHA-SUBUNIT OF ESCHERICHIA-COLI RNA-POLYMERASE - AMINO-ACID SUBSTITUTION WITHIN THE AMINO-TERMINAL ASSEMBLY DOMAIN [J].
KIMURA, M ;
ISHIHAMA, A .
JOURNAL OF MOLECULAR BIOLOGY, 1995, 254 (03) :342-349