Enzymatic Synthesis of Biologically Active Macromolecule

Maitra, Umadas

Abstract

The basic goal of our research program is to gain a fuller understanding of the molecular mechanisms of transcription and of initiation of translation of mRNA and the control of these processes in normal and viral infected cells.
The specific aims of the research proposed are directed toward the elucidation of the following major areas of interest: (1) Further characterization of a relatively simple DNA-dependent RNA polymerase (T3 RNA polymerase) which is induced in Escherichia coli cells following bacteriophage T3 infection, will be carried out particularly with regard to the various regulatory elements that control specific initiation and termination of RNA synthesis by T3 RNA polymerase from a T3 DNA template. The following areas are being pursued in detail: (a) characterization of promoters utilized in vitro by T3 RNA polymerase on T3 DNA; (b) identification of T3 RNA polymerase start sited in vivo in T3-infected cells in order to ascertain whether the same start sites are utilized by the phage polymerase in vivo as in vitro; (c) transcription of early genes of T3 genome by T3 RNA polymerase; (d) mechanism of transcription termination by T3 RNA polymerase. (2) Our laboratory is also concerned with identifying the eukaryotic initiation factors required for formation of functional 80S polypeptide chain initiation complexes, and in studying the interaction of these protein factors with ribosomes, Met-tRNAf, GTP and mRNA during initiation complex formation. The following areas are being actively pursued: (a) mechanism of catalytic reutilization of eukaryotic initiation factor 2 (eIF-2) in initiation complex formation; (b) further characterization of the reaction leading to the joining of 60S ribosomal subunits to 40S initiation complexes to form functional 80S initiation complexes; (c) protein factor requirements for natural mRNA-directed 40S initiation complex formation.