Protein surface engineering and interaction studies of maltogenic amylase towards improved enzyme immobilisation

Jaafar N. R., Jailani N., Rahman R. A., TOKSOY ÖNER E., Murad A. M. A., Illias R. M.

INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES, cilt.213, ss.70-82, 2022 (SCI-Expanded) identifier identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 213
  • Basım Tarihi: 2022
  • Doi Numarası: 10.1016/j.ijbiomac.2022.05.169
  • Dergi Adı: INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus, Academic Search Premier, BIOSIS, Biotechnology Research Abstracts, CAB Abstracts, Chemical Abstracts Core, EMBASE, Food Science & Technology Abstracts, INSPEC, MEDLINE, Veterinary Science Database
  • Sayfa Sayıları: ss.70-82
  • Anahtar Kelimeler: Computational analysis, Protein engineering, Enzyme immobilisation, Protein orientation, Protein secondary structure, HETEROFUNCTIONAL CARRIER, COVALENT IMMOBILIZATION, CANDIDA-RUGOSA, STABILITY, LIPASE, ORIENTATION, STRATEGIES, SUPPORTS, SITE, BENTONITE
  • Marmara Üniversitesi Adresli: Evet

Özet

A combined strategy of computational, protein engineering and cross-linked enzyme aggregates (CLEAs) approaches was performed on Bacillus lehensis G1 maltogenic amylase (Mag1) to investigate the preferred amino acids and orientation of the cross-linker in constructing stable and efficient biocatalyst. From the computational analysis, Mag1 exhibited the highest binding affinity towards chitosan (-7.5 kcal/mol) and favours having interactions with aspartic acid whereas glutaraldehyde was the least favoured (-3.4 kcal/mol) and has preferences for lysine. A total of eight Mag1 variants were constructed with either Asp or Lys substitutions on different secondary structures surface. Mutant Mag1-mDh exhibited the highest recovery activity (82.3%) in comparison to other Mag1 variants. Mutants-CLEAs exhibited higher thermal stability (20-30% activity) at 80 C whilst Mag1-CLEAs could only retain 9% of activity at the same temperature. Reusability analysis revealed that mutants-CLEAs can be recovered up to 8 cycles whereas Mag1-CLEAs activity could only be retained for up to 6 cycles. Thus, it is evident that amino acids on the enzyme's surface play a crucial role in the construction of highly stable, efficient and recyclable CLEAs. This demonstrates the necessity to determine the preferential amino acid by the cross-linkers in advance to facilitate CLEAs immobilisation for designing efficient biocatalysts.