Exploring the Structural and Dynamic Properties of a Chimeric Glycoside Hydrolase Protein in the Presence of Calcium Ions

Autor(es)

dos Santos, Alberto M. ; da Costa, Clauber H. S. ; Martins, Manoela ; Goldbeck, Rosana ; Skaf, Munir S.

• Tipo de documento: Artigo Científico
• Fonte: INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES; v. 25, n. 22, p. 16-pg., 2024-11-01.

Resumo

GH10 xylanases and GH62 Arabinofuranosidases play key roles in the breakdown of arabinoxylans and are important tools in various industrial and biotechnological processes, such as renewable biofuel production, the paper industry, and the production of short-chain xylooligosaccharides (XOS) from plant biomass. However, the use of these enzymes in industrial settings is often limited due to their relatively low thermostability and reduced catalytic efficiency. To overcome these limitations, strategies based on enzymatic chimera construction and the use of metal ions and other cofactors have been proposed to produce new recombinant enzymes with improved catalytic activity and thermostability. Here, we examine the conformational dynamics of a GH10-GH62 chimera at different calcium ion concentrations through molecular dynamics simulations. While experimental data have demonstrated improved activity and thermostability in GH10-GH62 chimera, the mechanistic basis for these enhancements remains unclear. We explored the structural details of the binding subsites of Ca2+ in the parental enzymes GH62 from Aspergillus fumigatus (Afafu62) and a recombinant GH10 from Cryptococcus flavescens (Xyn10cf), as well as their chimeric combination, and how negatively charged electron pairing located at the protein surface affects Ca2+ capture. The results indicate that Ca2+ binding significantly contributes to structural stability and catalytic cavity modulation in the chimera, particularly evident at a concentration of 0.01 M. This effect, not observed in the parental GH10 and GH62 enzymes, highlights how Ca2+ enhances stability in the overall chimeric enzyme, while supporting a larger cavity volume in the chimera GH62 subunit. The increased catalytic site volume and reduced structural flexibility in response to Ca2+ suggest that calcium binding minimizes non-productive conformational states, which could potentially improve catalytic turnover. The findings presented here may aid in the development of more thermostable and efficient catalytic systems. (AU)

• Processo FAPESP: 22/04695-2 - Mecanismos de reação de enzimas e aprimoramento catalítico com base em estados de transição e algoritmos de aprendizado de máquina
• Beneficiário: Alberto Monteiro dos Santos
• Modalidade de apoio: Bolsas no Brasil - Pós-Doutorado
• Processo FAPESP: 20/02871-2 - Proteína quimérica bifuncional de endoxilanase/arabinofuranosidase para hidrólise de resíduos agroindustriais e produção de xilo-oligossacarídeos
• Beneficiário: Manoela Martins
• Modalidade de apoio: Bolsas no Brasil - Doutorado
• Processo FAPESP: 22/04703-5 - Simulações de dinâmica molecular de enzimas de interesse biotecnológico
• Beneficiário: Clauber Henrique Souza da Costa
• Modalidade de apoio: Bolsas no Brasil - Pós-Doutorado
• Processo FAPESP: 13/08293-7 - CECC - Centro de Engenharia e Ciências Computacionais
• Beneficiário: Munir Salomao Skaf
• Modalidade de apoio: Auxílio à Pesquisa - Centros de Pesquisa, Inovação e Difusão - CEPIDs
• Processo FAPESP: 19/17874-0 - EMU concedido no Proc. 2013/08293-7, KAHUNA upgrade - HPE Apollo Gen10 Supercomputer
• Beneficiário: Munir Salomao Skaf
• Modalidade de apoio: Auxílio à Pesquisa - Programa Equipamentos Multiusuários