Structural model and functional properties of an exo-polygalacturonase from Neosartorya glabra

Author(s)

Desagiacomo, Carla Cristina Villela ^[1] ; Alnoch, Robson Carlos ^[2] ; Pinheiro, Vanessa Elisa ^[1] ; Cereia, Mariana ^[2] ; Machado, Carla Botelho ^[3] ; Damasio, Andre ^[4] ; Augusto, Marlei Josiele ^[5] ; Pedersoli, Wellington ^[1] ; Silva, Roberto Nascimento ^[1] ; Polizeli, Maria de Lourdes Teixeira de Moraes ^{[2, 1]}

Affiliation

^[1] Univ Sao Paulo, Fac Med Ribeirao Preto, Dept Bioquim & Imunol, BR-14049900 Ribeirao Preto, SP - Brazil

^[2] Univ Sao Paulo, Fac Filosofia Ciencias & Letras Ribeirao Preto, Dept Biol, BR-14040901 Ribeirao Preto, SP - Brazil

^[3] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol, BR-13083970 Campinas - Brazil

^[4] Univ Campinas UNICAMP, Inst Biol, BR-13083862 Campinas, SP - Brazil

^[5] Univ Sao Paulo, Fac Med Ribeirao Preto, Dept Patol & Med Legal, BR-14040901 Ribeirao Preto, SP - Brazil

• Document type: Journal article
• Source: International Journal of Biological Macromolecules; v. 186, p. 909-918, SEP 1 2021.
• Web of Science Citations: 0

Abstract

A purified exo-polygalacturonase of Neosartorya glabra (EplNg) was successfully characterized. EplNg native presented 68.2 kDa, with 32% carbohydrate content. The deglycosylated form showed 46.3 kDa and isoelectric point of 5.4. The identity of EpIlNg was confirmed as an exo-polygalacturonase class I (EC 3.2.1.67) using mass spectrometry and Western-Blotting. Capillary electrophoresis indicated that only galactumnic acid was released by the action of EplNg on sodium polypectate, confirming an exoenzyme character. The structural model confers that EplNg has a core formed by twisted parallel beta-sheets structure. Among twelve putative cysteines, ten were predicted to form disulfide bridges. The catalytic triad predicted is composed of Asp(223), ASp(245), and Asp(246) aligned along with a distance in 4-5 angstrom, suggesting that EplNg probably does not perform the standard inverting catalytic mechanism described for the GH28 family. EplNg was active from 30 to 90 degrees C, with maximum activity at 65 degrees C, pH 5.0. The Km and Vmax determined using sodium polypectate were 6.9 mg.mL(-1) and Vmax 690 mu mol.min(-1).mg(-1), respectively. EplNg was active and stable over a wide range of pH values and temperatures, confirming the interesting properties EplNg and provide a basis for the development of the enzyme in different biotechnological processes. (AU)

• FAPESP's process: 14/50884-5 - INCT 2014: National Institute of Science and Technology of Bioethanol
• Grantee: Marcos Silveira Buckeridge
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 20/00081-4 - Development of strategies for immobilization and co-immobilization of enzymes involved in biomass degradation
• Grantee: Robson Carlos Alnoch
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 18/07522-6 - Development of eco-friendly technologies for the hydrolysis of biomass and residues of cellulose pulp and paper industry
• Grantee: Maria de Lourdes Teixeira de Moraes Polizeli
• Support Opportunities: Regular Research Grants
• FAPESP's process: 17/22669-0 - N-glycosylation and enzymes secretion in filamentous fungi
• Grantee: André Ricardo de Lima Damasio
• Support Opportunities: Program for Research on Bioenergy (BIOEN) - Regular Program Grants