B.A. Chemistry Cornell University 1975
Ribonucleases, RNA Processing, and Gene Regulation
Ribonucleases are a functionally and mechanistically diverse group of enzymes that carry out essential RNA cleavage reactions involved in RNA maturation, RNA degradation, antiviral defense; and gene silencing. Ribonucleases are under intensive investigation in many laboratories, with the goal to understand their roles in gene expression and cell function in both the normal and disease states. Also, specifically engineered ribonucleases are being developed as potent anticancer agents. My research group is specifically interested in the ribonuclease III (RNase III) superfamily of double-stranded(ds)-RNA-specific endonucleases, that are highly conserved in prokaryotic and eukaryotic cells. We are studying primarily the mechanism of action of bacterial RNase III orthologs, which exhibit a conserved dsRNA-binding domain (dsRBD) and a nuclease domain. The crystal structure of RNase III of Thermotoga maritima is shown in the figure, which exhibits a two-fold symmetric (a)2 subunit structure, with the extended dsRBD "arms." We are characterizing the active site components of Escherichia coli RNase III, and determining divalent metal ion function in phosphodiester hydrolysis. We also are characterizing the sequence and structural features in RNase III substrates that confer reactivity and cleavage site selectivity. We have determined that specific Watson-Crick base-pair sequences act as antideterminants (i.e. inhibitors) of RNase III recognition and cleavage. These sequence elements appear to play a vital role in cleavage site selection as well as in protecting other intracellular dsRNAs from inappropriate cleavage. Current research is extending the findings on E. coli RNase III to thermophilic bacterial orthologs, including Aquifex aeolicus RNase III and Thermotoga maritima RNase III.
Alla, N.R.; Nicholson A.W. 2012. "Evidence for a dual functional role of a conserved histidine in RNA.DNA heteroduplex cleavage by human RNase H1." Febs Journal. 279(23): 4492-4500
Gone, S.; Nicholson, A.W. 2012. "Bacteriophage 17 protein kinase: Site of inhibitory autophosphorylation, and use of dephosphorylated enzyme for efficient modification of protein in vitro." Protein Expression and Purification. 85(2): 218-223
Nicholson, A.W. 2012. "Dissecting Human Dicer: Some Assembly Required." Journal of Molecular Biology. 422(4): 464-465
Shi, Z.J.; Nicholson, R.H.; Jaggi, R.; Nicholson, A.W.; 2011 "Characterization of Aquifex aeolicus ribonuclease III and the reactivity epitopes of its pre-ribosomal RNA substrates." Nucleic Acids Research. 39(7): 2756-2768
Nathania, L.; Nicholson, A.W. 2010. "Thermotoga maritima Ribonuclease III. Characterization of Thermostable Biochemical Behavior and Analysis of Conserved Base Pairs That Function as Reactivity Eoptopes for the Thermotoga 23S rRNA Precursor." Biochemistry. 49(33): 7164-7178
Meng, W.Z.; Nicholson, R.H.; Nathania, L.; Nicholson, A.W. 2008. "New Approaches to Understanding Double-Stranded RNA Processing By Ribonuclease III: Purification and Assays of Homodimeric and Heterodimerica Forms of RNase III from Bacterial Extremophiles and Mesophiles." RNA Turnover in Bacteria, Archaea and Organelles. 447: 119-129
Meng, W.; Nicholson, A.W. 2008. "Heterodimer-based analysis of subunit and domain contributions to double-stranded RNA processing by Escherichia coli RNase III in vitro." Biochem J. 410: 39-48.