B.A. Chemistry Cornell University 1975 Ph.D. Chemistry University of Pennsylvania 1981 NIH Postdoctoral Fellow Rockefeller University 1981-85

Biological Chemistry

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.

Pertzev, AV.; Nicholson, A.W. 2006. "Characterization of RNA sequence determinants and antideterminants of processing reactivity for a minimal substrate of Escherichia coli ribonuuclease III." Nucleic Acids Res. 34: 3708-3721.

Sun, W., Pertzev, A., Nicholson, A.W. 2005. "Catalytic mechanism of Escherichia coli ribonuclease III. Kinetic and inhibitor evidence for the involvement of two magnesium ions in RNA phosphodiester hydrolysis." Nucleic Acids Res. 33: 807-815.

Sun, W., Li, G. and Nicholson, A.W. 2004. "Mutational analysis of the nuclease domain of Escherichia coli ribonuclease III. Identification of conserved residues that are important for catalytic function in vitro." Biochem. 43: 13054-13062.

Zhang, Y., Calin-Jageman, I., Gurnon, J.R., Choi, T.J., Adams, B., Nicholson, A.W. and Van Etten, J.L. 2003. "Characterization of a Chlorella virus PBCV-1 encoded ribonuclease III. Virol. 317: 73-83.

Nicholson, A. W. 2003. "The ribonuclease III superfamily: forms and functions in RNA maturation, decay, and gene silencing." In: "RNAi: A Guide to Gene Silencing" (G. Hannon, Ed) Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY .