%0 Journal Article %1 WOS:000305366100040 %A Silva, Fredy D A %A Vasconcelos, Ilka M %A Lobo, Marina D P %A de Castro, Patricia G %A Magalhaes, Vladimir G %A de Freitas, Cleverson D T %A Carlini, Celia R R S %A Pinto, Paulo M %A Beltramini, Leila M %A Filho, Jose H A %A Barros, Eduardo B %A Alencar, Luciana M R %A Grangeiro, Thalles B %A Oliveira, Jose T A %C RADARWEG 29, 1043 NX AMSTERDAM, NETHERLANDS %D 2012 %I ELSEVIER %J BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS %K Amino Circular Vigna acid cDNA dichroism; sequence; sequence} unguiculata; {2-Cys-peroxiredoxin; %N 7 %P 1128-1140 %R 10.1016/j.bbagen.2012.03.003 %T Biochemical, physicochemical and molecular characterization of a genuine 2-Cys-peroxiredoxin purified from cowpea Vigna unguiculata (L.) Walpers leaves %V 1820 %X Background: Peroxiredoxins have diverse functions in cellular defense-signaling pathways. 2-Cys-peroxiredoxins (2-Cys-Prx) reduce H2O2 and alkyl-hydroperoxide. This study describes the purification and characterization of a genuine 2-Cys-Prx from Vigna unguiculata (Vu-2-Cys-Prx). Methods: Vu-2-Cys-Prx was purified from leaves by ammonium sulfate fractionation, chitin affinity and ion exchange chromatography. Results: Vu-2-Cys-Prx reduces H2O2 using NADPH and DTT. Vu-2-Cys-Prx is a 44 kDa (SDS-PAGE)/46 kDa (exclusion chromatography) protein that appears as a 22 kDa molecule under reducing conditions, indicating that it is a homodimer linked intermolecularly by disulfide bonds and has a pI range of 4.56-4.72; its NH2-terminal sequence was similar to 2-Cys-Prx from Phaseolus vulgaris (96%) and Populus tricocarpa (96%). Analysis by ESI-Q-TOF MS/MS showed a molecular mass/pI of 28.622 kDa/5.18. Vu-2-Cys-Prx has 8% alpha-helix, 39% beta-sheet, 22% of turns and 31% of unordered forms. Vu-2-Cys-Prx was heat stable, has optimal activity at pH 7.0, and prevented plasmid DNA degradation. Atomic force microscopy shows that Vu-2-Cys-Prx oligomerized in decamers which might be associated with its molecular chaperone activity that prevented denaturation of insulin and citrate synthase. Its cDNA analysis showed that the redox-active Cys(52) residue and the amino acids Pro(45), Thr(49) and Arg(128) are conserved as in other 2-Cys-Prx. General significance: The biochemical and molecular features of Vu-2-Cys-Prx are similar to other members of 2-Cys-Prx family. To date, only one publication reported on the purification of native 2-Cys-Prx from leaves and the subsequent analysis by N-terminal Edman sequencing, which is crucial for construction of stromal recombinant 2-Cys-Prx proteins. (C) 2012 Elsevier B.V. All rights reserved.

@article{WOS:000305366100040, abstract = {Background: Peroxiredoxins have diverse functions in cellular defense-signaling pathways. 2-Cys-peroxiredoxins (2-Cys-Prx) reduce H2O2 and alkyl-hydroperoxide. This study describes the purification and characterization of a genuine 2-Cys-Prx from Vigna unguiculata (Vu-2-Cys-Prx). Methods: Vu-2-Cys-Prx was purified from leaves by ammonium sulfate fractionation, chitin affinity and ion exchange chromatography. Results: Vu-2-Cys-Prx reduces H2O2 using NADPH and DTT. Vu-2-Cys-Prx is a 44 kDa (SDS-PAGE)/46 kDa (exclusion chromatography) protein that appears as a 22 kDa molecule under reducing conditions, indicating that it is a homodimer linked intermolecularly by disulfide bonds and has a pI range of 4.56-4.72; its NH2-terminal sequence was similar to 2-Cys-Prx from Phaseolus vulgaris (96%) and Populus tricocarpa (96%). Analysis by ESI-Q-TOF MS/MS showed a molecular mass/pI of 28.622 kDa/5.18. Vu-2-Cys-Prx has 8% alpha-helix, 39% beta-sheet, 22% of turns and 31% of unordered forms. Vu-2-Cys-Prx was heat stable, has optimal activity at pH 7.0, and prevented plasmid DNA degradation. Atomic force microscopy shows that Vu-2-Cys-Prx oligomerized in decamers which might be associated with its molecular chaperone activity that prevented denaturation of insulin and citrate synthase. Its cDNA analysis showed that the redox-active Cys(52) residue and the amino acids Pro(45), Thr(49) and Arg(128) are conserved as in other 2-Cys-Prx. General significance: The biochemical and molecular features of Vu-2-Cys-Prx are similar to other members of 2-Cys-Prx family. To date, only one publication reported on the purification of native 2-Cys-Prx from leaves and the subsequent analysis by N-terminal Edman sequencing, which is crucial for construction of stromal recombinant 2-Cys-Prx proteins. (C) 2012 Elsevier B.V. All rights reserved.}, added-at = {2022-05-23T20:00:14.000+0200}, address = {RADARWEG 29, 1043 NX AMSTERDAM, NETHERLANDS}, author = {Silva, Fredy D A and Vasconcelos, Ilka M and Lobo, Marina D P and de Castro, Patricia G and Magalhaes, Vladimir G and de Freitas, Cleverson D T and Carlini, Celia R R S and Pinto, Paulo M and Beltramini, Leila M and Filho, Jose H A and Barros, Eduardo B and Alencar, Luciana M R and Grangeiro, Thalles B and Oliveira, Jose T A}, biburl = {https://www.bibsonomy.org/bibtex/2e7223925a182d8371e7aca3e6ead0b97/ppgfis_ufc_br}, doi = {10.1016/j.bbagen.2012.03.003}, interhash = {4615064750da35920e38de97fc005792}, intrahash = {e7223925a182d8371e7aca3e6ead0b97}, issn = {0304-4165}, journal = {BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS}, keywords = {Amino Circular Vigna acid cDNA dichroism; sequence; sequence} unguiculata; {2-Cys-peroxiredoxin;}, number = 7, pages = {1128-1140}, publisher = {ELSEVIER}, pubstate = {published}, timestamp = {2022-05-23T20:00:14.000+0200}, title = {Biochemical, physicochemical and molecular characterization of a genuine 2-Cys-peroxiredoxin purified from cowpea [Vigna unguiculata (L.) Walpers] leaves}, tppubtype = {article}, volume = 1820, year = 2012 }