A newly synthesized, ribosome-bound polypeptide chain adopts conformations dissimilar from early in vitro refolding intermediates

被引:71
作者
Clark, PL [1 ]
King, J [1 ]
机构
[1] MIT, Dept Biol 68 340D, Cambridge, MA 02139 USA
关键词
D O I
10.1074/jbc.M008490200
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
Little is known about the conformations of newly synthesized polypeptide chains as they emerge from the large ribosomal subunit, or how these conformations compare with those populated immediately after dilution of polypeptide chains out of denaturant in vitro. Both in vivo and in vitro, partially folded intermediates of the tailspike protein from Salmonella typhimurium phage P22 can be trapped in the cold. A subset of monoclonal antibodies raised against tailspike recognize partially folded intermediates, whereas other antibodies recognize only later intermediates and/or the native state. We have used a pair of monoclonal antibodies to probe the conformational features of full-length, newly synthesized tailspike chains recovered on ribosomes from phage-infected cells. The antibody that recognizes early intermediates in vitro also recognizes the ribosome-bound intermediates. Surprisingly, the antibody that did not recognize early in vitro intermediates did recognize ribosome-bound tailspike chains translated in vivo, Thus, the newly synthesized, ribosome-bound tailspike chains display structured epitopes not detected upon dilution of tailspike chains from denaturant. As opposed to the random ensemble first populated when polypeptide chains are diluted out of denaturant, folding in vivo from the ribosome may begin with polypeptide conformations already directed toward the productive folding and assembly pathway.
引用
收藏
页码:25411 / 25420
页数:10
相关论文
共 65 条
[1]   PRINCIPLES THAT GOVERN FOLDING OF PROTEIN CHAINS [J].
ANFINSEN, CB .
SCIENCE, 1973, 181 (4096) :223-230
[2]   The complete atomic structure of the large ribosomal subunit at 2.4 Å resolution [J].
Ban, N ;
Nissen, P ;
Hansen, J ;
Moore, PB ;
Steitz, TA .
SCIENCE, 2000, 289 (5481) :905-920
[3]   PURIFICATION AND ORGANIZATION OF THE GENE-1 PORTAL PROTEIN REQUIRED FOR PHAGE-P22 DNA PACKAGING [J].
BAZINET, C ;
BENBASAT, J ;
KING, J ;
CARAZO, JM ;
CARRASCOSA, JL .
BIOCHEMISTRY, 1988, 27 (06) :1849-1856
[4]   NASCENT POLYPEPTIDE-CHAINS EMERGE FROM THE EXIT DOMAIN OF THE LARGE RIBOSOMAL-SUBUNIT - IMMUNE MAPPING OF THE NASCENT CHAIN [J].
BERNABEU, C ;
LAKE, JA .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES, 1982, 79 (10) :3111-3115
[5]   Cold rescue of the thermolabile tailspike intermediate at the junction between productive folding and off-pathway aggregation [J].
Betts, SD ;
King, J .
PROTEIN SCIENCE, 1998, 7 (07) :1516-1523
[6]   MECHANISM OF HEAD ASSEMBLY AND DNA ENCAPSULATION IN SALMONELLA PHAGE-P22 .1. GENES, PROTEINS, STRUCTURES AND DNA MATURATION [J].
BOTSTEIN, D ;
WADDELL, CH ;
KING, J .
JOURNAL OF MOLECULAR BIOLOGY, 1973, 80 (04) :669-695
[7]  
BRUNSCHIER R, 1993, J BIOL CHEM, V268, P2767
[8]   Chaperone properties of bacterial elongation factor EF-G and initiation factor IF2 [J].
Caldas, T ;
Laalami, S ;
Richarme, G .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2000, 275 (02) :855-860
[9]  
CAMPBELL AM, 1996, ESCHERICHIA COLI SAL, P2325
[10]   X-ray crystal structures of 70S ribosome functional complexes [J].
Cate, JH ;
Yusupov, MM ;
Yusupova, GZ ;
Earnest, TN ;
Noller, HF .
SCIENCE, 1999, 285 (5436) :2095-2104