Kettering University

About: Kettering University is a education organization based out in Flint, Michigan, United States. It is known for research contribution in the topics: RNA & Antigen. The organization has 6842 authors who have published 7689 publications receiving 337503 citations. The organization is also known as: GMI Engineering & Management Institute & General Motors Institute.

Structure of an Rrp6–RNA exosome complex bound to poly(A) RNA

Abstract: The eukaryotic RNA exosome processes and degrades RNA by directing substrates to the distributive or processive 3′ to 5′ exoribonuclease activities of Rrp6 or Rrp44, respectively. The non-catalytic nine-subunit exosome core (Exo9) features a prominent central channel. Although RNA can pass through the channel to engage Rrp44, it is not clear how RNA is directed to Rrp6 or whether Rrp6 uses the central channel. Here we report a 3.3 A crystal structure of a ten-subunit RNA exosome complex from Saccharomyces cerevisiae composed of the Exo9 core and Rrp6 bound to single-stranded poly(A) RNA. The Rrp6 catalytic domain rests on top of the Exo9 S1/KH ring above the central channel, the RNA 3′ end is anchored in the Rrp6 active site, and the remaining RNA traverses the S1/KH ring in an opposite orientation to that observed in a structure of a Rrp44-containing exosome complex. Solution studies with human and yeast RNA exosome complexes suggest that the RNA path to Rrp6 is conserved and dependent on the integrity of the S1/KH ring. Although path selection to Rrp6 or Rrp44 is stochastic in vitro, the fate of a particular RNA may be determined in vivo by the manner in which cofactors present RNA to the RNA exosome. The exosome complex contains two catalytic subunits which degrade RNA in either a distributive (Rrp6) or a processive (Rrp44) manner—previous structures indicated how RNA could be directed to Rrp44, but the path taken to Rrp6 was unclear; here the location of the Rrp6 catalytic domain and the RNA 3′ end are determined and it is found that the RNA lies in an opposite orientation from that of the Rrp44-containing exosome structure, suggesting that the fate of an RNA may be influenced by the manner in which cofactors present it. The exosome complex contains two catalytic subunits that degrade RNA in either a distributive (Rrp6) or a processive (Rrp44) manner. Previous structures indicated how RNA could be directed to Rrp44, but the path taken to Rrp6 was unclear. Christopher Lima and colleagues have now solved the crystal structure of a yeast ten-subunit exosome complex (lacking Rrp44) bound to single-stranded RNA. The location of the Rrp6 catalytic domain and the RNA 3' end are determined, but more importantly, it is found that the RNA lies in an orientation opposite that from the Rrp44-containing exosome structure. This result suggests that the fate of an RNA may be a stochastic decision based on how the RNA binds to the complex.

Recombination mediated by vaccinia virus DNA topoisomerase I in Escherichia coli is sequence specific

Abstract: Specialized type I topoisomerases catalyze DNA strand transfer during site-specific recombination in prokaryotes and fungi. As a rule, the site specificity of these systems is determined by the DNA binding and cleavage preference of the topoisomerase per se. The Mr 32,000 topoisomerase I encoded by vaccinia virus (a member of the eukaryotic family of "general" type I enzymes) is also selective in its interaction with DNA; binding and cleavage occur in vitro at a pentameric motif 5'-(C or T)CCTT in duplex DNA. Expression of vaccinia virus DNA topoisomerase I in a lambda lysogen of Escherichia coli promotes int-independent excisive recombination of the prophage. To address whether the topoisomerase directly catalyzes DNA strand transfer in vivo, the recombination junctions of plaque-purified progeny phage were cloned and sequenced. In five of six distinct excision events examined, a topoisomerase cleavage sequence is present in one strand of the DNA duplex of both recombining partners. Recombination entails no duplication, insertion, or deletion of nucleotides at the crossover points, consistent with excision via conservative strand exchange at sites of topoisomerase cleavage. Three of these five recombination events are distinguished by the presence of direct repeats at the parental half-sites that extend beyond the pentameric cleavage motif, suggesting that sequence homology may facilitate excision. The data are consistent with a model in which vaccinia topoisomerase catalyzes reciprocal strand transfer, leading to the formation of a nonmigrating Holliday junction, the resolution of which can lead to excisive recombination.

Mitomycin c: chemical and biological studies on alkylation.

Abstract: The presence of an aziridine ring in mitomycin C suggests that the mechanism of action of the antibiotic is like that of the antitumor alkylating agents. However the compound is unexpectedly stable during aerobic incubation with rat liver homogenates although rapidly metabolized anaerobically. Mitomycin is not reactive with gamma-(4-nitrobenzyl) pyridine and reacts only slowly at acid p(H) with thiosulfate. It is proposed that mitomycin is activated in vivo, possibly by a reduction which "unmasks" the potential activity of the fused aziridine ring.