Isolation and Characterization of Functional Ribosomal RNA Fragments

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; R o o t E n t r y F C o m p O b j b W o r d D o c u m e n t O b j e c t P o o l /Q /Q 1 2 3 4 5 6 7 8 9 : ; F Microsoft Word 6.0 Document MSWordDoc Word.Document.6 ; V t C; r 9^ u F ^ ~ t ~ u F v j v F )F ~ tT ~ tNj;j V P " V P G # V F j v F P O ~ u hd V Pj @ # 3 v v j V ' v j j v V Pj Vj 2# ~ v! v N )F ~ v Wh $ 'Nu v v v $9420768 Mankin The focus of this research is the function of different regions of ribosomal RNA. A new approach for identification of functionally significant rRNA segments is proposed. Namely, an expression library of random rRNA fragments will be screened for RNA species which can affect ribosome functions in vivo. The efficiency of the approach has been tested in preliminary studies where an active rRNA fragment conferring resistance to the ribosome targeted antibiotic erythromycin has been isolated from a random library of rRNA fragments. Analysis of the function of the erythromycin resistance RNA is one of the aims of the research. Further, functional selection protocols will be used to identify clones which express other active rRNA fragments that 1) alter sensitivity to ribosome targeted antibiotics, 2) interfere with ribosome assembly and rRNA processing, 3) affect accuracy of translation and 4) interfere with protein biosynthesis. Genetic and biochemical analysis will be used to determine structural requirements for biological activity of the selected rRNA fragments and the mode of their a ction. This should lead to a better understanding of the functions of corresponding rRNA regions in the ribosome. %%% All proteins in the cell are synthesized by ribosomes. The ribosome is composed of ribosomal RNA and protein molecules. Function of the ribosomal RNA is the subject of the proposed research. A new approach will be used in the proposed research. Ribosomal RNA will be cut into small random pieces. Functional fragments will be isolated and their function studied. An attempt will be made to identify fragments of the ribosomal RNA that can influence assembly of the ribosome, accuracy of protein synthesis and sensitivity to antibiotics. In general, the proposed approach should lead to a better understanding of the functions of rRNA in the process of protein biosynthesis. *** @ ....()()))()() S u m m a r y I n f o r m a t i o n ( 0 Oh +' 0 $ H l D h R:\WWUSER\TEMPLATE\NORMAL.DOT 9420768 marcia steinberg marcia steinberg @ C @ @ C @ Microsoft Word 6.0 2 ; e 3 e y C 0 ! T u ; h 9420768 Mankin The focus of this research is the function of different regions of ribosomal RNA. A new approach for identification of functionally significant rRNA segments is proposed. Namely, an expression library of random rRNA fragments will be screened for RNA species which can affect ribosome functions in vivo. The efficiency of the approach has been tested in preliminary studies where an active rRNA fragment conferring resistance to the ribosome targeted antibiotic erythromycin has been isolated from a random library of rRNA fragments. Analysis of the functio