@article{cobucci-ponzano_zorzetti_strazzulli_carillo_bedini_corsaro_comfort_kelly_rossi_moracci_2011, title={A novel alpha-d-galactosynthase from Thermotoga maritima converts beta-d-galactopyranosyl azide to alpha-galacto-oligosaccharides}, volume={21}, ISSN={["1460-2423"]}, DOI={10.1093/glycob/cwq177}, abstractNote={The large-scale production of oligosaccharides is a daunting task, hampering the study of the role of glycans in vivo and the testing of the efficacy of novel glycan-based drugs. Glycosynthases, mutated glycosidases that synthesize oligosaccharides in high yields, are becoming important chemo-enzymatic tools for the production of oligosaccharides. However, while β-glycosynthase can be produced with a rather well-established technology, examples of α-glycosynthases are thus far limited only to enzymes from glycoside hydrolase 29 (GH29), GH31 and GH95 families. α-L-Fucosynthases from GH29 use convenient glycosyl azide derivatives as a strategic alternative to glycosyl fluoride donors. However, the general applicability of this method to other α-glycosynthases is not trivial and remains to be confirmed. Here, β-D-galactopyranosyl azide was converted to α-galacto-oligosaccharides with good yields and high regioselectivity, catalyzed by a novel α-galactosynthase based on the GH36 α-galactosidase from the hyperthermophilic bacterium Thermotoga maritima. These results open a new avenue to the practical synthesis of biologically interesting α-galacto-oligosaccharides and demonstrate more widespread use of β-glycosyl-azide as donors, confirming their utility to expand the repertoire of glycosynthases.}, number={4}, journal={GLYCOBIOLOGY}, author={Cobucci-Ponzano, Beatrice and Zorzetti, Carmela and Strazzulli, Andrea and Carillo, Sara and Bedini, Emiliano and Corsaro, Maria Michela and Comfort, Donald A. and Kelly, Robert M. and Rossi, Mose and Moracci, Marco}, year={2011}, month={Apr}, pages={448–456} }
 @article{comfort_chou_conners_vanfossen_kelly_2008, title={Functional-genomics-based identification and characterization of open reading frames encoding alpha-glucoside-processing enzymes in the hyperthermophilic archaeon Pyrococcus furiosus}, volume={74}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.01920-07}, abstractNote={ABSTRACT}, number={4}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Comfort, Donald A. and Chou, Chung-Jung and Conners, Shannon B. and VanFossen, Amy L. and Kelly, Robert M.}, year={2008}, month={Feb}, pages={1281–1283} }
 @article{comfort_bobrov_ivanen_shabalin_harris_kulminskaya_brumer_kelly_2007, title={Biochemical analysis of Thermotoga maritima GH36 alpha-galactosidase (TmGalA) confirms the mechanistic commonality of clan GH-D glycoside hydrolases}, volume={46}, ISSN={["0006-2960"]}, DOI={10.1021/bi061521n}, abstractNote={Organization of glycoside hydrolase (GH) families into clans expands the utility of information on catalytic mechanisms of member enzymes. This issue was examined for GH27 and GH36 through biochemical analysis of GH36 alpha-galactosidase from Thermotoga maritima (TmGalA). Catalytic residues in TmGalA were inferred through structural homology with GH27 members to facilitate design of site-directed mutants. Product analysis confirmed that the wild type (WT) acted with retention of anomeric stereochemistry, analogous to GH27 enzymes. Conserved acidic residues were confirmed through kinetic analysis of D327G and D387G mutant enzymes, azide rescue, and determination of azide rescue products. Mutation of Asp327 to Gly resulted in a mutant that had a 200-800-fold lower catalytic rate on aryl galactosides relative to the WT enzyme. Azide rescue experiments using the D327G enzyme showed a 30-fold higher catalytic rate compared to without azide. Addition of azide to the reaction resulted in formation of azide beta-d-galactopyranoside, confirming Asp327 as the nucleophilic residue. The Asp387Gly mutation was 1500-fold catalytically slower than the WT enzyme on p-nitrophenyl alpha-d-galactopyranoside. Analysis at different pH values produced a bell-shaped curve of the WT enzyme, but D387G exhibited higher activity with increasing pH. Catalyzed reactions with the D387G mutant in the presence of azide resulted in formation of azide alpha-d-galactopryanoside as the product of a retaining mechanism. These results confirm that Asp387 is the acid/base residue of TmGalA. Furthermore, they show that the biochemical characteristics of GH36 TmGalA are closely related to GH27 enzymes, confirming the mechanistic commonality of clan GH-D members.}, number={11}, journal={BIOCHEMISTRY}, author={Comfort, Donald A. and Bobrov, Kirill S. and Ivanen, Dina R. and Shabalin, Konstantin A. and Harris, James M. and Kulminskaya, Anna A. and Brumer, Harry and Kelly, Robert M.}, year={2007}, month={Mar}, pages={3319–3330} }
 @article{chou_shockley_conners_lewis_comfort_adams_kelly_2007, title={Impact of substrate glycoside linkage. and elemental sulfur on bioenergetics, of and hydrogen production by the hyperthermophilic Archaeon Pyrococcus furiosus}, volume={73}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.00597-07}, abstractNote={ABSTRACT}, number={21}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Chou, Chung-Jung and Shockley, Keith R. and Conners, Shannon B. and Lewis, Derrick L. and Comfort, Donald A. and Adams, Michael W. W. and Kelly, Robert M.}, year={2007}, month={Nov}, pages={6842–6853} }
 @article{mahammad_comfort_kelly_khan_2007, title={Rheological properties of guar galactomannan solutions during hydrolysis with galactomannanase and alpha-galactosidase enzyme mixtures}, volume={8}, ISSN={["1526-4602"]}, DOI={10.1021/bm0608232}, abstractNote={Guar galactomannan, a naturally occurring polysaccharide, is susceptible to hydrolysis by three enzymes: beta-mannosidase, beta-mannanase, and alpha-galactosidase. The beta-mannosidase cleaves a single mannose unit from the nonreducing end of the guar molecule, the beta-mannanase cleaves interior glycosidic bonds between adjacent mannose units, and the alpha-galactosidase cleaves the galactose side branches off the guar. In this study, hydrolysis of guar solutions using hyperthermopilic versions of these enzymes together in different proportions and combinations are examined. The enzymatic reactions are carried out in situ in a rheometer, and the progress of the reaction is monitored through measuring the variation in zero shear viscosity. We find the presence of alpha-galactosidase to affect the action of both beta-mannanase and beta-mannosidase with respect to solution rheology. However, this effect is more pronounced when the alpha-galactosidase and beta-mannanase or beta-mannosidase enzymes were added sequentially rather than simultaneously. This likely is the result of debranching of the guar, which facilitates attack on beta-1,4-linkages by both the beta-mannanase and the beta-mannosidase enzymes and increases hydrolytic rates by the individual enzymes. A rheology-based kinetic model is developed to estimate the reaction rate constants and interpret synergistic effects of multiple enzyme contributions. The model fits the experimental data well and reveals that both the native and the debranched guar have the same activation energy for beta-mannanase action, although debranching considerably increases the frequency of enzyme-guar interactions.}, number={3}, journal={BIOMACROMOLECULES}, author={Mahammad, Shamsheer and Comfort, Donald A. and Kelly, Robert M. and Khan, Saad A.}, year={2007}, month={Mar}, pages={949–956} }
 @article{lee_shockley_schut_conners_montero_johnson_chou_bridger_wigner_brehm_et al._2006, title={Transcriptional and biochemical analysis of starch metabolism in the hyperthermophilic archaeon Pyrococcus furiosus}, volume={188}, ISSN={["1098-5530"]}, DOI={10.1128/JB.188.6.2115-2125.2006}, abstractNote={ABSTRACT}, number={6}, journal={JOURNAL OF BACTERIOLOGY}, author={Lee, HS and Shockley, KR and Schut, GJ and Conners, SB and Montero, CI and Johnson, MR and Chou, CJ and Bridger, SL and Wigner, N and Brehm, SD and et al.}, year={2006}, month={Mar}, pages={2115–2125} }
 @article{conners_montero_comfort_shockley_johnson_chhabra_kelly_2005, title={An expression-driven approach to the prediction of carbohydrate transport and utilization regulons in the hyperthermophilic bacterium Thermotoga maritima}, volume={187}, ISSN={["1098-5530"]}, DOI={10.1128/JB.187.21.7267-7282.2005}, abstractNote={ABSTRACT}, number={21}, journal={JOURNAL OF BACTERIOLOGY}, author={Conners, SB and Montero, CI and Comfort, DA and Shockley, KR and Johnson, MR and Chhabra, SR and Kelly, RM}, year={2005}, month={Nov}, pages={7267–7282} }