ctbe/lnbio: publications in partnership
TRANSCRIPT
Partnership with LNBio
The Brazilian Bioethanol Science and Technology Laboratory (CTBE) publications printed in partnership with Brazilian Biosciences National Laboratory (LNBio) and indexed by Web of Sciences (www.isiwebofknowledge.com) are listed below in chronological order.
Criteria for articles selectionFill “(LNBIO OR Lab Nacl Biociencias) and (CTBE or bioetanol or bioethanol) and Brazil” in the Address Search form.
Total of CTBE/LNBio Journal Articles11.
11. Functional characterization and oligomerization of a recombinant xyloglucan-specific endo-beta-1,4-glucanase (GH12)from Aspergillus niveus
Author(s): Damasio, ARL; Ribeiro, LFC; Ribeiro, LF; Furtado, GP; Segato, F; Almeida, FBR; Crivellari, AC; Buckeridge, MS; Souza, TACB; Murakami, MT; Ward, RJ; Prade, RA ; Polizeli, MLTM | E-mail: [email protected] Source: BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS Volume: 1824 Issue: 3 Pages: 461-467 | DOI: 10.1016/j.bbapap.2011.12.005 | Published: MAR 2012
Abstract: Xyloglucan is a major structural polysaccharide of the primary (growing) cell wall of higher plants. It consists of a cellulosic backbone (beta-1,4-linked glucosyl residues) that is frequently substituted with side chains. This report describes Aspergillus nidulans strain A773 recombinant secretion of a dimeric xyloglucan-specific endo-beta-1,4-glucanohydrolase (XegA) cloned from Aspergillus niveus. The ORF of the A. niveus xegA gene is comprised of 714 nucleotides, and encodes a 238 amino acid protein with a calculated molecular weight of 23.5 kDa and isoelectric point of 4.38. The optimal pH and temperature were 6.0 and 60 degrees C, respectively. XegA generated a xyloglucan-oligosaccharides (XGOs) pattern similar to that observed for cellulases from family GH12, i.e., demonstrating that its mode of action includes hydrolysis of the glycosidiclinkages between glucosyl residues that are not branched with xylose. In contrast to commercial lichenase, mixed
linkagebeta-glucan (lichenan) was not digested by XegA, indicating that the enzyme did not cleave glucan beta-1,3 or beta-1,6 bonds.The far-UV CD spectrum of the purified enzyme indicated a protein rich in beta-sheet structures as expected for GH12 xyloglucanases. Thermal unfolding studies displayed two transitions with mid-point temperatures of 51.3 degrees C and 81.3degrees C respectively, and dynamic light scattering studies indicated that the first transition involves a change in oligomeric state from a dimeric to a monomeric form. Since the enzyme is a predominantly a monomer at 60 degrees C. the enzymatic assays demonstrated that XegA is more active in its monomeric state.Full Text: www.sciencedirect.com/science/article/pii/S1570963911003220
Addresses: [Polizeli, Maria L. T. M.] Univ Sao Paulo, Fac Filosofia Ciencias & Letras Ribeirao Preto, Dept Biol, BR-14040901 Ribeirao Preto, SP, Brazil [Damasio, Andre R. L.; Ribeiro, Liliane F. C.; Ribeiro, Lucas F.; Furtado, Gilvan P.] Univ Sao Paulo, Dept Bioquim & Imunol, FMRP, BR-14040901 Ribeirao Preto, SP, Brazil[Segato, Fernando; Buckeridge, Marcos S.] Ctr Nacl Pesquisas Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, Campinas, SP, Brazil[Almeida, Fausto B. R.] Univ Sao Paulo, Dept Biol Celular & Mol & Bioagentes Patogen, FMRP, BR-14040901 Ribeirao Preto, SP, Brazil[Crivellari, Augusto C.; Buckeridge, Marcos S.] Univ Sao Paulo, Inst Biociencias, Dept Bot, Lab Fisiol Ecol Plantas, BR-14040901 Ribeirao Preto, SP, Brazil[Souza, Tatiana A. C. B.; Murakami, Mario T.] Ctr Nacl Pesquisas Energia & Mat, Lab Nacl Biociencias, Campinas, SP, Brazil[Ward, Richard J.] Univ Sao Paulo, Dept Quim, FFCLRP USP, BR-14040901 Ribeirao Preto, SP, Brazil[Prade, Rolf A.] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA
10. Molecular insights into substrate specificity and thermal stability of a bacterial GH5-CBM27 endo-1,4-beta-Dmannanase
Author(s): dos Santos, CR; Paiva, JH; Meza, AN; Cota, J; Alvarez, TM; Ruller, R; Prade, RA; Squina, FM; Murakami, MT | E-mail: [email protected]: JOURNAL OF STRUCTURAL BIOLOGY Volume: 177 Issue: 2 Pages: 469-476 | DOI:10.1016/j.jsb.2011.11.021 | Published: FEB 2012
Abstract: The breakdown of beta-1,4-mannoside linkages in a variety of mannan-containing polysaccharides is of great importance in industrial processes such as kraft pulp delignification, food processing and production of second-generation biofuels, which puts a premium on studies regarding the prospection and engineering of beta-mannanases. In this work, a two-domain beta-mannanase from Thermotoga petrophila that encompasses a GH5 catalytic domain with a C-terminal CBM27 accessory domain, was functionally and structurally characterized. Kinetic and thermal denaturation experiments showed that the CBM27 domain provided thermo-protection to the catalytic domain, while no contribution on enzymatic activity was observed. The structure of the catalytic domain determined by SIRAS revealed a canonical (alpha/beta)(8)-barrel scaffold surrounded by loops and short helices that form the catalytic interface. Several structurally related ligand molecules interacting with TpMan were solved at high-resolution and resulted in a wide-range representation of the subsites forming the active-site cleft with residues W134, E198, R200, E235, H283 and W284 directly involved in glucose binding.Full Text: www.sciencedirect.com/science/article/pii/S1047847711003376
Addresses: [dos Santos, Camila Ramos; Paiva, Joice Helena; Meza, Andreia Navarro; Murakami, Mario Tyago] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias LNBio, BR-13083970 Campinas, SP, Brazil
[Cota, Junio; Alvarez, Thabata Maria; Ruller, Roberto; Squina, Fabio Marcio] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, BR-13083970 Campinas, SP, Brazil[Prade, Rolf Alexander] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA
9. Dissecting structure-function-stability relationships of a thermostable GH5-CBM3 cellulase from Bacillus subtilis 168
Author(s): Santos, CR; Paiva, JH; Sforca, ML; Neves, JL; Navarro, RZ; Cota, J; Akao, PK; Hoffmam, ZB ; Meza, AN; Smetana, JH; Nogueira, ML; Polikarpov, I; Xavier-Neto, J; Squina, FM; Ward, RJ; Ruller, R; Zeri, AC; Murakami, MT | E-mail: [email protected] Source: BIOCHEMICAL JOURNAL Volume: 441 Pages: 95-104 | DOI: 10.1042/BJ20110869 Part: Part 1 | Published: JAN 1 2012
Abstract: Cellulases participate in a number of biological events, such as plant cell wall remodelling, nematode parasitism and microbial carbon uptake. Their ability to depolymerize crystalline cellulose is of great biotechnological interest for environmentally compatible production of fuels from lignocellulosic biomass. However, industrial use of cellulases is somewhat limited by both their low catalytic efficiency and stability. In the present study, we conducted a detailed functional and structural characterization of the thermostable BsCe15A (Bacillus subtilis cellulase 5A), which consists of a GH5 (glycoside hydrolase 5) catalytic domain fused to a CBM3 (family 3 carbohydrate-binding module). NMR structural analysis revealed that the Bacillus CBM3 represents a new subfamily, which lacks the classical calcium-binding motif, and variations in NMR frequencies in the presence of cellopentaose showed the importance of polar residues in the carbohydrate interaction. Together with the catalytic domain, the CBM3 forms a large planar surface for cellulose recognition, which conducts the substrate in a proper conformation to the active site and increases enzymatic efficiency. Notably, the manganese ion wasdemonstrated to have a hyper-stabilizing effect on BsCel5A, and by using deletion constructs and X-ray crystallography we determined that this effect maps to a negatively charged motif located at the opposite face of the catalytic site.Full Text: www.biochemj.org/bj/441/bj4410095.htm
Addresses: [Santos, Camila R.; Paiva, Joice H.; Sforca, Mauricio L.; Neves, Jorge L.; Navarro, Rodrigo Z.; Akao, Patricia K.; Meza, Andreia N.; Smetana, Juliana H.; Nogueira, Maria L.; Xavier-Neto, Jose; Zeri, Ana C.; Murakami, Mario T.] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias, Campinas, SP, Brazil[Cota, Junio; Hoffmam, Zaira B.; Squina, Fabio M.; Ruller, Roberto] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol, Campinas, SP, Brazil[Polikarpov, Igor] Univ Sao Paulo, Inst Fis Sao Carlos, Sao Carlos, SP, Brazil[Ward, Richard J.] Univ Sao Paulo, Dept Quim, BR-14049 Ribeirao Preto, Brazil
8. Dissecting structure-function-stability relationships of a thermostable GH5-CBM3 cellulase from Bacillus subtilis 168
Author(s): Santos, CR; Paiva, JH; Sforca, ML; Neves, JL; Navarro, RZ; Cota, J; Akao, PK; Hoffmam, ZB ; Meza, AN; Smetana, JH; Nogueira, ML; Polikarpov, I; Xavier-Neto, J; Squina, FM; Ward, RJ; Ruller, R; Zeri, AC; Murakami, MT | E-mail: [email protected] Source: BIOCHEMICAL JOURNAL Volume: 441 Pages: 95-104 | DOI: 10.1042/BJ20110869 Part: Part 1 | Published: JAN 1 2012
Abstract: Cellulases participate in a number of biological events, such as plant cell wall remodelling, nematode parasitism and microbial carbon uptake. Their ability to depolymerize crystalline cellulose is of great biotechnological interest for environmentally compatible production of fuels from
lignocellulosic biomass. However, industrial use of cellulases is somewhat limited by both their low catalytic efficiency and stability. In the present study, we conducted a detailed functional and structural characterization of the thermostable BsCe15A (Bacillus subtilis cellulase 5A), which consists of a GH5 (glycoside hydrolase 5) catalytic domain fused to a CBM3 (family 3 carbohydrate-binding module). NMR structural analysis revealed that the Bacillus CBM3 represents a new subfamily, which lacks the classical calcium-binding motif, and variations in NMR frequencies in the presence of cellopentaose showed the importance of polar residues in the carbohydrate interaction. Together with the catalytic domain, the CBM3 forms a large planar surface for cellulose recognition, which conducts the substrate in a proper conformation to the active site and increases enzymatic efficiency. Notably, the manganese ion wasdemonstrated to have a hyper-stabilizing effect on BsCel5A, and by using deletion constructs and X-ray crystallography we determined that this effect maps to a negatively charged motif located at the opposite face of the catalytic site.Full Text: www.biochemj.org/bj/441/bj4410095.htm
Addresses: [Santos, Camila R.; Paiva, Joice H.; Sforca, Mauricio L.; Neves, Jorge L.; Navarro, Rodrigo Z.; Akao, Patricia K.; Meza, Andreia N.; Smetana, Juliana H.; Nogueira, Maria L.; Xavier-Neto, Jose; Zeri, Ana C.; Murakami, Mario T.] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias, Campinas, SP, Brazil[Cota, Junio; Hoffmam, Zaira B.; Squina, Fabio M.; Ruller, Roberto] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol, Campinas, SP, Brazil[Polikarpov, Igor] Univ Sao Paulo, Inst Fis Sao Carlos, Sao Carlos, SP, Brazil[Ward, Richard J.] Univ Sao Paulo, Dept Quim, BR-14049 Ribeirao Preto, Brazil
7. Structure of a novel thermostable GH51 alpha-L-arabinofuranosidase from Thermotoga petrophila RKU-1
Author(s): Souza, TACB; Santos, CR; Souza, AR; Oldiges, DP; Ruller, R; Prade, RA; Squina, FM; Murakami, MT | E-mail: [email protected] Source: PROTEIN SCIENCE Volume: 20 Issue: 9 Pages: 1632-1637 | DOI: 10.1002/pro.693 | Published: SEP 2011
Abstract: alpha-L-arabinofuranosidases (EC 3.2.1.55) participate in the degradation of a variety of L-arabinose-containing polysaccharides and interact synergistically with other hemicellulases in the production of oligosaccharides and bioconversion of lignocellulosic biomass into biofuels. In this work, the structure of a novel thermostable family 51 (GH51) alpha-L-arabinofuranosidase from Thermotoga petrophila RKU-1 (TpAraF) was determined at 3.1 angstrom resolution. The TpAraF tertiary structure consists of an (alpha/beta)-barrel catalytic core associated with a C-terminal beta-sandwich domain, which is stabilized by hydrophobic contacts. In contrast to other structurally characterized GH51 AraFs, the accessory domain of TpAraF is intimately linked to the active site by a long beta-hairpin motif, which modifies the catalytic cavity in shape and volume. Sequence and structural analyses indicate that this motif is unique to Thermotoga AraFs. Small angle X-ray scattering investigation showed that TpAraF assembles as a hexamer in solution and is preserved at the optimum catalytic temperature, 65 degrees C, suggesting functional significance. Crystal packing analysis shows that the biological hexamer encompasses a dimer of trimers and the multiple oligomeric interfaces are predominantly fashioned by polar and electrostatic contacts.Full Text: http://onlinelibrary.wiley.com/doi/10.1002/pro.693/full
Addresses: [Souza, TACB; Santos, CR; Souza, AR; Oldiges, DP; Murakami, MT] Ctr Nacl Pesquisa Energia & Mat, LNBio, BR-13083970 Campinas, SP, Brazil[Ruller, R; Squina, FM] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia Tecnol Bioetanol CTBE, BR-13083970 Campinas, SP, Brazil[Prade, RA] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA
6. Heterologous expression of an Aspergillus niveus xylanase GH11 in Aspergillus nidulans and its characterization and application
Author(s): Damasio, ARD; Silva, TM; Almeida, FBD; Squina, FM; Ribeiro, DA; Leme, AFP; Segato, F;Prade, RA; Jorge, JA; Terenzi, HF; Polizeli, MDTM | E-mail: [email protected] Source: PROCESS BIOCHEMISTRY Volume: 46 Issue: 6 Pages: 1236-1242 | DOI:10.1016/j.procbio.2011.01.027 | Published: JUN 2011
Abstract: A xylanase was cloned from Aspergillus niveus and successfully expressed in Aspergillus nidulans (XAN). The full-length gene consisted of 890 bp and encoded 275 mature amino acids with a calculated mass of 31.3 kDa. The deduced amino acid sequence was highly homologous with the xylanase belonging to family 11 of the glycoside hydrolases. The recombinant protein was purified to electrophoretic homogeneity by anion-exchange chromatography and gel filtration. The optima of pH and temperature for the recombinant enzyme were 5.0 and 65 degrees C, respectively. The thermal stability of the recombinant xylanase was extremely improved by covalent immobilization on glyoxyl agarose with 91.4% of residual activity after 180 min at 60 degrees C, on the other hand, the free xylanase showed a half-life of 9.9 min at the same temperature.Affinity chromatography on Concanavalin A- and Jacalin-agarose columns followed by SDS-PAGE analyses showed that the XAN has O- and N-glycans. XAN promotes hydrolysis of xylan resulting in xylobiose, xylotriose and xylotetraose.Intermediate degradation of xylan resulting in xylo-oligomers is appealing for functional foods as the beneficial effect of oligosaccharides on gastrointestinal micro flora includes preventing proliferation of pathogenic intestinal bacteria and facilitates digestion and absorption of nutrients.Full Text: www.sciencedirect.com/science/article/pii/S135951131100033X
Addresses: [Silva, TM; Jorge, JA; Terenzi, HF; Polizeli, MDTM] Univ Sao Paulo, Fac Filosofia Ciencias & Letras Ribeirao Preto, Dept Biol, BR-14040901 Ribeirao Preto, SP, Brazil[Damasio, ARD] Univ Sao Paulo, Fac Med Ribeirao Preto, Dept Bioquim & Imunol, BR-14040901 Ribeirao Preto, SP, Brazil[Almeida, FBD] Univ Sao Paulo, Fac Med Ribeirao Preto, Dept Biol Celular & Mol & Bioagentes Patogen, BR-14040901 Ribeirao Preto, SP, Brazil[Squina, FM; Ribeiro, DA] Ctr Nacl Pesquisa Energia & Mat CNPEM, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, Campinas, SP, Brazil[Leme, AFP] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias LNBio, Campinas, SP, Brazil[Segato, F; Prade, RA] Oklahoma State Univ, Dept Mol Biol & Microbiol, Stillwater, OK 74078 USA
5. Mode of operation and low-resolution structure of a multi-domain and hyperthermophilic endo-beta-1,3-glucanase from Thermotoga petrophila
Author(s): Cota, J; Alvarez, TM; Citadini, AP; Santos, CR; Neto, MD; Oliveira, RR; Pastore, GM; Ruller, R; Prade, RA; Murakami, MT; Squina, FM | E-mail: [email protected]: BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Volume: 406 Issue: 4 Pages: 590-594 | DOI: 10.1016/j.bbrc.2011.02.098 | Published: MAR 25 2011
Abstract: 1,3-beta-Glucan depolymerizing enzymes have considerable biotechnological applications including biofuel production, feedstock-chemicals and pharmaceuticals. Here we describe a comprehensive functional characterization and low-resolution structure of a hyperthermophilic laminarinase from Thermotoga petrophila (TpLam). We determine TpLam enzymatic mode of operation, which specifically cleaves internal beta-1,3-glucosidic bonds. The enzyme most frequently attacks the bond between the 3rd and 4th residue from the non-reducing end, producing glucose,
laminaribiose and laminaritriose as major products. Far-UV circular dichroism demonstrates that TpLam is formed mainly by beta structural elements, and the secondary structure is maintained after incubation at 90 degrees C. The structure resolved by small angle X-ray scattering, reveals a multi-domain structural architecture of a V-shape envelope with a catalytic domain flanked by two carbohydrate-binding modules. Full Text: www.sciencedirect.com/science/article/pii/S0006291X11003032
Addresses: [Cota, J; Alvarez, TM; Citadini, AP; Ruller, R; Squina, FM] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, Campinas, SP, Brazil[Santos, CR; Oliveira, RR; Murakami, MT] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias LNBio, Campinas, SP, Brazil[Neto, MD] Univ Sao Paulo, Inst Fis Sao Carlos IFSC, Sao Carlos, SP, Brazil[Cota, J; Pastore, GM] Univ Estadual Campinas, Fac Engn Alimentos FEA, Campinas, SP, Brazil[Prade, RA] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA
4. Functional and biophysical characterization of a hyperthermostable GH51 alpha-L-arabinofuranosidase from Thermotoga petrophila
Author(s): dos Santos, CR; Squina, FM; Navarro, AM; Oldiges, DP; Leme, AFP; Ruller, R; Mort, AJ; Prade, R; Murakami, MT | E-mail: [email protected]: BIOTECHNOLOGY LETTERS Volume: 33 Issue: 1 Pages: 131-137 | DOI:10.1007/s10529-010-0409-3 | Published: JAN 2011
Abstract: A hyperthermostable glycoside hydrolase family 51 (GH51) alpha-L-arabinofuranosidase from Thermotoga petrophila RKU-1 (TpAraF) was cloned, overexpressed, purified and characterized. The recombinant enzyme had optimum activity at pH 6.0 and 70 degrees C with linear alpha-1,5-linked arabinoheptaose as substrate. The substrate cleavage pattern monitored by capillary zone electrophoresis showed that TpAraF is a classical exo-acting enzyme producing arabinose as its end-product. Far-UV circular dichroism analysis displayed a typical spectrum of alpha/beta barrel proteins analogously observed for other GH51 alpha-L-arabinofuranosidases. Moreover, TpAraF was crystallized in two crystalline forms, which can be used to determine its crystallographic structure.Full Text: www.springerlink.com/content/w27417057jr58l17/
Addresses: [dos Santos, CR; Navarro, AM; Oldiges, DP; Leme, AFP; Murakami, MT] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias LNBio, BR-13083970 Campinas, SP, Brazil[Squina, FM; Ruller, R] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, BR-13083970 Campinas, SP, Brazil[Mort, AJ] Oklahoma State Univ, Dept Biochem, Stillwater, OK 74078 USA[Prade, R] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA
3. Thermal-induced conformational changes in the product release area drive the enzymatic activity of xylanases 10B: Crystal structure, conformational stability and functional characterization of the xylanase 10B from Thermotoga petrophila RKU-1
Author(s): Santos, CR; Meza, AN; Hoffmam, ZB; Silva, JC; Alvarez, TM; Ruller, R; Giesel, GM; Verli, H; Squina, FM; Prade, RA; Murakami, MT | E-mail: [email protected]: BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Volume: 403 Issue: 2 Pages: 214-219 | DOI: 10.1016/j.bbrc.2010.11.010 | Published: DEC 10 2010
Abstract: Endo-xylanases play a key role in the depolymerization of xylan and recently, they have attracted much attention owing to their potential applications on biofuels and paper industries. In this
work, we have investigated the molecular basis for the action mode of xylanases 10B at high temperatures using biochemical, biophysical and crystallographic methods. The crystal structure of xylanase 10B from hyperthermophilic bacterium Thermotoga petrophila RKU-1 (TpXyl10B) has been solved in the native state and in complex with xylobiose. The complex crystal structure showed a classical binding mode shared among other xylanases, which encompasses the -1 and -2 subsites. Interestingly, TpXyl10B displayed a temperaturedependent action mode producing xylobiose and xylotriose at 20 degrees C, and exclusively xylobiose at 90 degrees C as assessed by capillary zone electrophoresis. Moreover, circular dichroism spectroscopy suggested a coupling effect of temperature-induced structural changes with this particular enzymatic behavior. Molecular dynamics simulations supported the CD analysis suggesting that an open conformational state adopted by the catalytic loop (Trp297-Lys326) provokes significant modifications in the product release area (+1,+2 and +3 subsites), which drives the enzymatic activity to the specific release of xylobiose at high temperatures. (C) 2010 Elsevier Inc. All rights reserved.Full Text: http://www.sciencedirect.com/science/article/pii/S0006291X10020589
Addresses: [Santos, CR; Meza, AN; Murakami, MT] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias LNBio, BR-13083970 Campinas, SP, Brazil[Hoffmam, ZB; Silva, JC; Alvarez, TM; Ruller, R; Squina, FM] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, BR-13083970 Campinas, SP, Brazil[Giesel, GM; Verli, H] Univ Fed Rio Grande do Sul, Ctr Biotecnol, Porto Alegre, RS, Brazil[Prade, RA] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA
2. Substrate cleavage pattern, biophysical characterization and low-resolution structure of a novel hyperthermostable arabinanase from Thermotoga petrophila
Author(s): Squina, FM; Santos, CR; Ribeiro, DA; Cota, J; de Oliveira, RR; Ruller, R; Mort, A; Murakami, MT; Prade, RA | E-mail: [email protected] Source: BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Volume: 399 Issue: 4 Pages: 505-511 | DOI: 10.1016/j.bbrc.2010.07.097 | Published: SEP 3 2010
Abstract: Arabinan is a plant structural polysaccharide degraded by two enzymes: alpha-L-arabinofuranosidase and endo-1,5-alpha-L-arabinanase. These enzymes are highly diversified in nature, however, little is known about their biochemical and biophysical properties. We have characterized a novel arabinanase (AbnA) isolated from Thermotoga petrophila with unique thermostable properties such as the insignificant decrease of residual activity after incubation up to 90 degrees C. We determined the AbnA mode of operation through capillary zone electrophoresis, which accumulates arabinotriose and arabinobiose as end products after hydrolysis of arabinan-containing polysaccharides. Spectroscopic analyses by Far-UV circular dichroism and intrinsic tryptophan fluorescence emission demonstrated that AbnA is folded and formed mainly by beta-sheet structural elements. In silico molecular modeling showed that the AbnA structure encompasses a five-bladed beta-propeller catalytic core juxtaposed by distorted up-and-down beta-barrel domain. The low-resolution structure determined by small angle X-ray scattering indicated that AbnA is monomeric in solution and its molecular shape is in full agreement with the model. (C) 2010 Elsevier Inc. All rights reserved.Full Text: http://www.sciencedirect.com/science/article/pii/S0006291X10014178
Addresses: [Squina, FM; Ribeiro, DA; Cota, J; Ruller, R] CNPEM, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, Campinas, SP, Brazil[Santos, CR; de Oliveira, RR; Murakami, MT] CNPEM, Lab Nacl Biociencias LNBio, Campinas, SP, Brazil [Cota, J] Univ Estadual Campinas, Dept Ciencias Alimentos, BR-13081970 Campinas, SP, Brazil[Mort, A] Oklahoma State Univ, Dept Biochem, Stillwater, OK 74078 USA
[Prade, RA] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA
1. Cloning, expression, purification, crystallization and preliminary X-ray diffraction studies of the catalytic domain of a hyperthermostable endo-1,4-beta-d-mannanase from Thermotoga petrophila RKU-1
Author(s): Santos, CR; Squina, FM; Navarro, AM; Ruller, R; Prade, R; Murakami, MT | E-mail: [email protected] Source: ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY AND CRYSTALLIZATION COMMUNICATIONS Volume: 66 Pages: 1078-1081 | DOI: 10.1107/S1744309110029131 | Published: SEP 2010
Abstract: Endo-1,4-beta-d-mannanases play key roles in seed germination and fruit ripening and have recently received much attention owing to their potential applications in the food, detergent and kraft pulp industries. In order to delineate their structural determinants for specificity and stability, X-ray crystallographic investigations combined with detailed functional studies are being performed. In this work, crystals of the catalytic domain of a hyperthermostable endo-1,4-beta-d-mannanase from Thermotoga petrophila RKU-1 were obtained from three different conditions, resulting in two crystalline forms. Crystals from conditions with phosphate or citrate salts as precipitant (CryP) belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 58.76, b = 87.99, c = 97.34 A, while a crystal from a condition with ethanol as precipitant (CryE) belonged to space group I2(1)2(1)2(1), with unit-cell parameters a = 91.03, b = 89.97, c = 97.89 A. CryP and CryE diffracted to resolutions of 1.40 and 1.45 A, respectively.Full Text: http://journals.iucr.org/f/issues/2010/09/00/hc5111/index.html
Addresses: [Santos, CR; Navarro, AM; Murakami, MT] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Biociencias LNBio, BR-13083970 Campinas, SP, Brazil[Squina, FM; Ruller, R] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol CTBE, BR-13083970 Campinas, SP, Brazil[Prade, R] Oklahoma State Univ, Dept Microbiol & Mol Genet, Stillwater, OK 74078 USA