Suzanne F. O'Handley
Rochester Institute of Technology
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Featured researches published by Suzanne F. O'Handley.
Journal of Biological Chemistry | 1996
Maurice J. Bessman; David N. Frick; Suzanne F. O'Handley
Our studies on the biochemical basis of spontaneous mutations took an interesting and unexpected turn when we discovered that a small region of amino acid homology between the MutT protein of Escherichia coli and the MutX protein of Streptococcus pneumoniae was involved in their nucleoside triphosphatase as well as their antimutator activities (1–3). Computer searches of the data banks revealed that this same small conserved region was present in a number of other proteins in organisms ranging from viruses to humans (2, 4). Most of these proteins containing the signature are coded for by open reading frames (orfs) whose products are either unidentified or of unknown function. We have been attempting, systematically, to identify and characterize enzymatic activities associated with these proteins, and it is now evident that nature has adopted this motif, originally identified as the active site of the nucleoside-triphosphate pyrophosphohydrolase of MutT (5, 6), and adapted it for use in many diverse reactions distinct from its function in the MutT protein. This short review summarizes our present knowledge of those reactions catalyzed by proteins harboring the MutT signature sequence and calls attention to a unique and versatile nucleotide binding and catalytic site. Although it might appear that the enzymes of this family act upon a wide variety of unrelated substrates, those characterized so far all hydrolyze a nucleoside diphosphate linked to some other moiety, X. For convenience, and to correct a misapprehension, we propose the mnemonic “nudix” hydrolase for this family of enzymes to replace the “MutT family.” Currently, this signature sequence is designated the “MutT pattern” in version 13.0 of the PROSITE data base of amino acid sequence motifs (7). This initial classification is misleading, because many, if not most, of these proteins are not involved directly in preventing mutations nor do they catalyze the archetypal nucleoside triphosphate pyrophosphohydrolysis reaction originally described for MutT itself (5, 6).
Journal of Biological Chemistry | 1999
Christopher A. Dunn; Suzanne F. O'Handley; David N. Frick; Maurice J. Bessman
Four Nudix hydrolase genes, ysa1 fromSaccharomyces cerevisiae, orf209 fromEscherichia coli, yqkg from Bacillus subtilis, and hi0398 from Hemophilus influenzae were amplified, cloned into an expression vector, and transformed into E. coli. The expressed proteins were purified and shown to belong to a subfamily of Nudix hydrolases active on ADP-ribose. Comparison with other members of the subfamily revealed a conserved proline 16 amino acid residues downstream of the Nudix box, common to all of the ADP-ribose pyrophosphatase subfamily. In this same region, a conserved tyrosine designates another subfamily, the diadenosine polyphosphate pyrophosphatases, while an array of eight conserved amino acids is indicative of the NADH pyrophosphatases. On the basis of these classifications, the trgB gene, a tellurite resistance factor from Rhodobacter sphaeroides, was predicted to designate an ADP-ribose pyrophosphatase. In support of this hypothesis, a highly specific ADP-ribose pyrophosphatase gene from the archaebacterium, Methanococcus jannaschii, introduced into E. coli, increased the transformants tolerance to potassium tellurite.
Journal of Biological Chemistry | 1998
Saifuddin Sheikh; Suzanne F. O'Handley; Christopher A. Dunn; Maurice J. Bessman
The MJ1149 gene from the Archaeon,Methanococcus jannaschii, has been cloned and expressed inEscherichia coli. The 19-kDa protein containing the Nudix box, GX5EX7REUXEEXGU, has been purified and identified as a highly specific enzyme catalyzing the Mg2+-dependent hydrolysis of ADP-ribose according to the equation: ADP-ribose + H2O → AMP + ribose-5-phosphate. The enzyme retains full activity when heated to 80 °C, and the rate of hydrolysis is 15-fold higher at 75 °C than at 37 °C in keeping with the thermophilicity of the organism. This is the first Nudix hydrolase identified from the Archaea, indicating that the family of enzymes containing the Nudix signature sequence is represented in all three kingdoms.
Structure | 2003
Lin Woo Kang; Sandra B. Gabelli; Jennifer E. Cunningham; Suzanne F. O'Handley; L. Mario Amzel
Nudix hydrolases are a family of proteins that contain the characteristic sequence GX(5)EX(7)REUXEEXG(I/L/V), the Nudix box. They catalyze the hydrolysis of a variety of nucleoside diphosphate derivatives such as ADP-ribose, Ap(n)A (3 </= n </= 6), NADH, and dATP. A number of Nudix hydrolases from several species, ranging from bacteria to humans, have been characterized, including, in some cases, the determination of their three-dimensional structures. The product of the Rv1700 gene of M. tuberculosis is a Nudix hydrolase specific for ADP-ribose (ADPR). We have determined the crystal structures of MT-ADPRase alone, and in complex with substrate, with substrate and the nonactivating metal ion Gd(3+), and in complex with a nonhydrolyzable ADPR analog and the activating metal ion Mn(2+). These structures, refined with data extending to resolutions between 2.0 and 2.3 A, showed that there are sequence differences in binding site residues between MT-ADPRase and a human homolog that may be exploited for antituberculosis drug development.
Journal of Biological Chemistry | 1996
Suzanne F. O'Handley; David N. Frick; L C Bullions; Albert S. Mildvan; Maurice J. Bessman
The product of the Escherichia coli orf17 gene is a novel nucleoside triphosphate pyrophosphohydrolase with a preference for dATP over the other canonical (deoxy)nucleoside triphosphates, and it catalyzes the hydrolysis of dATP through a nucleophilic attack at the β-phosphorus to produce dAMP and inorganic pyrophosphate. It has a pH optimum between 8.5 and 9.0, a divalent metal ion requirement with optimal activity at 5 mM Mg2+, a Km of 0.8 mM and a kcat of 5.2 s−1 at 37°C for dATP. dAMP is a weak competitive inhibitor with a Ki of approximately 4 mM, while PPi is a much stronger inhibitor with an apparent Ki of approximately 20 μM. The enzyme contains the highly conserved signature sequence GXVEX2ETX6REVXEEX2I designating the MutT family of proteins. However, unlike the other nucleoside triphosphate pyrophosphohydrolases with this conserved sequence, the Orf17 protein does not complement the mutT− mutator phenotype, and thus must serve a different biological role in the cell.
Journal of Biological Chemistry | 2006
WenLian Xu; Christopher A. Dunn; Suzanne F. O'Handley; Denise L. Smith; Maurice J. Bessman
Three members of the Nudix (nucleoside diphosphate X) hydrolase superfamily have been cloned from Escherichia coli MG1655 and expressed. The proteins have been purified and identified as enzymes active on nucleoside diphosphate derivatives with the following specificities. Orf141 (yfaO) is a nucleoside triphosphatase preferring pyrimidine deoxynucleoside triphosphates. Orf153 (ymfB) is a nonspecific nucleoside tri- and diphosphatase and atypically releases inorganic orthophosphate from triphosphates instead of pyrophosphate. Orf191 (yffH) is a highly active GDP-mannose pyrophosphatase. All three enzymes require a divalent cation for activity and are optimally active at alkaline pH, characteristic of the Nudix hydrolase superfamily. The question of whether or not Orf1.9 (wcaH) is a bona fide member of the Nudix hydrolase superfamily is discussed.
Biochemical Journal | 2006
Isaac M. Tirrell; Jennifer L. Wall; Christopher J. Daley; Sarah J. Denial; Frances G. Tennis; Kevin G. Galens; Suzanne F. O'Handley
YZGD from Paenibacillus thiaminolyticus is a novel bifunctional enzyme with both PLPase (pyridoxal phosphatase) and Nudix (nucleoside diphosphate x) hydrolase activities. The PLPase activity is catalysed by the HAD (haloacid dehalogenase) superfamily motif of the enzyme, and the Nudix hydrolase activity is catalysed by the conserved Nudix signature sequence within a separate portion of the enzyme, as confirmed by site-directed mutagenesis. YZGDs phosphatase activity is very specific, with pyridoxal phosphate being the only natural substrate, while YZGDs Nudix activity is just the opposite, with YZGD being the most versatile Nudix hydrolase characterized to date. YZGDs Nudix substrates include the CDP-alcohols (CDP-ethanol, CDP-choline and CDP-glycerol), the ADP-coenzymes (NADH, NAD and FAD), ADP-sugars, TDP-glucose and, to a lesser extent, UDP- and GDP-sugars. Regardless of the Nudix substrate, one of the products is always a nucleoside monophosphate, suggesting a role in nucleotide salvage. Both the PLPase and Nudix hydrolase activities require a bivalent metal cation, but while PLPase activity is supported by Co2+, Mg2+, Zn2+ and Mn2+, the Nudix hydrolase activity is Mn2+-specific. YZGDs phosphatase activity is optimal at an acidic pH (pH 5), while YZGDs Nudix activities are optimal at an alkaline pH (pH 8.5). YZGD is the first enzyme reported to be a member of both the HAD and Nudix hydrolase superfamilies, the first PLPase to be recognized as a member of the HAD superfamily and the first Nudix hydrolase capable of hydrolysing ADP-x, CDP-x and TDP-x substrates with comparable substrate specificity.
Science | 1992
Che Fu Kuo; Duncan E. McRee; Cindy L. Fisher; Suzanne F. O'Handley; Richard P. Cunningham; John A. Tainer
Biochemistry | 1993
Suzanne F. O'Handley; David G. Sanford; Rong Xu; Cathy C. Lester; Brian E. Hingerty; Suse Broyde; Thomas R. Krugh
Journal of Biological Chemistry | 1998
Suzanne F. O'Handley; David N. Frick; Christopher A. Dunn; Maurice J. Bessman