Regulation of the B. Subtilis trp Genes by the trp RNA- binding Attenuation Protein (TRAP)

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9603594 Gollnick The importance of RNA binding proteins as regulators of gene expression has become increasingly clear in recent years. The trp genes of Bacillus subtilis are regulated by an RNA binding protein called TRAP (trp RNA-binding Attenuation Protein). TRAP regulates the expression of trp genes in three ways which all involve tryptophan-dependent binding of TRAP to RNA. Transcription of the trpEDCFBA operon is controlled by an attenuation mechanism. TRAP binding to a site containing 11 (G/U)AG repeats in the leader sequence of trp operon transcript affects the secondary structure of the attenuator and promotes formation of a transcription terminator. TRAP also regulates translation of the trpE and trpG genes. TRAP is proposed to sequester the ribosome binding site of the trpE mRNA by altering the secondary structure of the mRNA leader sequence, and to compete directly with ribosomes for binding to the trpG mRNA. Recent X-ray crystallographic analysis of the TRAP-L-tryptophan complex revealed a novel macromolecule consisting of 11 identical subunits arranged in ring structure with a 23 Angstrom hole. Each subunit is composed entirely of beta-strands, beta-turns and random coils. The discoveries that TRAP is an 11-subunit protein and that its trp RNA binding site contains multiple small sequence repeats has led to a model in which the trp RNA wraps around the TRAP so that each of the 11 repeats in the trp RNA leader sequence interacts with one TRAP subunit or a combination of adjacent subunits. The objective of this research is to determine precisely how TRAP recognizes its target RNAs including the identification of the RNA binding domain of TRAP and the determinants of target RNA specificity for TRAP, and the characterization of the kinetics and thermodynamics of TRAP - RNA interaction. This research will provide new information about how proteins recognize RNA and will result in better understanding of the basic principles of protein-RNA interaction that may govern more com plicated processes of transcription, translation, RNA processing and transport in eukaryotes. The importance of RNA binding proteins as regulators of gene expression has become increasingly clear in recent years. The genes responsible for tryptophan biosynthesis in the bacteria Bacillus subtilis are regulated by an RNA binding protein called TRAP (trp RNA-binding Attenuation Protein), which requires the amino acid tryptophan to bind to RNA. Recent structural analysis indicates that TRAP is a novel macromolecule consisting of 11 identical subunits arranged in ring structure. This research will determine how TRAP recognizes and binds to its target RNA, and will provide new information and a better understanding of the basic principles of protein-RNA interaction that may govern more complicated processes of transcription, translation, RNA processing and transport in eukaryotes.