Kelley W L, Georgopoulos C
Departement de Biochimie Médicale, Centre Médical Universitaire, Université de Genève, Geneva, Switzerland.
Proc Natl Acad Sci U S A. 1997 Apr 15;94(8):3679-84. doi: 10.1073/pnas.94.8.3679.
The N-terminal 70 residue "J-domain" of the Escherichia coli DnaJ molecular chaperone is the defining and highly conserved feature of a large protein family. Based upon limited, yet significant, amino acid sequence homology to the J-domain, the DNA encoding the T/t common exon of the simian virus 40 (SV40), JC, or BK polyoma virus T antigen oncoproteins was used to construct J-domain replacement chimeras of the E. coli DnaJ chaperone. The virally encoded J-domains successfully substituted for the bacterial counterpart in vivo as shown by (i) complementation for viability at low and high temperature of a hypersensitive bacterial reporter strain, and (ii) the restoration of bacteriophage lambda plaque forming ability in the same strain. The amino acid change, H42Q, in the SV40 T/t and the JC virus T/t exon, which is positionally equivalent to the canonical dnaJ259 H33Q mutation within the E. coli J-domain, entirely abolished complementing activity. These results strongly suggest that the heretofore functionally undefined viral T/t common exon represents a bona fide J-domain that preserves critical features of the characteristic domain fold essential for J-domain interaction with the ATPase domain of the Hsp70 family. This finding has implications for the regulation of DNA tumor virus T antigens by molecular chaperones.
大肠杆菌DnaJ分子伴侣的N端70个残基的“J结构域”是一个大型蛋白质家族的决定性且高度保守的特征。基于与J结构域有限但显著的氨基酸序列同源性,编码猿猴病毒40(SV40)、JC或BK多瘤病毒T抗原癌蛋白T/t共同外显子的DNA被用于构建大肠杆菌DnaJ伴侣的J结构域替代嵌合体。如(i)对超敏细菌报告菌株在低温和高温下的生存能力进行互补,以及(ii)在同一菌株中恢复噬菌体λ的噬菌斑形成能力所示,病毒编码的J结构域在体内成功替代了细菌对应物。SV40 T/t和JC病毒T/t外显子中的氨基酸变化H42Q,其位置与大肠杆菌J结构域内的典型dnaJ259 H33Q突变相当,完全消除了互补活性。这些结果强烈表明,迄今为止功能未明的病毒T/t共同外显子代表一个真正的J结构域,它保留了对于J结构域与Hsp70家族ATP酶结构域相互作用至关重要的特征结构域折叠的关键特征。这一发现对分子伴侣对DNA肿瘤病毒T抗原的调控具有重要意义。