Mutational analysis of flavonol synthase of M. pinnata towards enhancement of binding affinity: a computational approach

Abstract

Millettia pinnata is an important medicinal plant that has been used as a treatment of various diseases due to presence of wide range of pharmacological properties. The plant contains quercetin, kaempferol, karanjin, pongaglabrone, kanjone, kanugin, gammatin, pongaglabol, and other bioflavonoids. Kaempferol is a natural flavonol that shows many pharmacological properties including anti-inflammatory, antioxidant, anticancer, and antidiabetic activities etc. The enzyme flavonol synthase (FLS, EC 1.14.20.6) catalyses the conversion of dihydroflavonols to flavonols, i.e. biosynthesis of kaempferol from dihydrokaempferol. The current work examined the binding affinity-based approach to improve the enzyme catalytic activity using computational methods. Sequential site-directed mutagenesis was used to create four mutants with the goal to increase hydrogen bonds and further improving the ligand (dihydrokaempferol) binding efficiency. Simulations were done to monitor the stability of the mutants followed by molecular docking to confirm interactions with ligand. For structure validation, various dynamic analysis like RMSD, RMSF, ROG, SASA, H-bond, PCA, DCCM, and FEL were performed, which predicts the stability of wild-type (WT) proteins and mutants. The Mutant_2 and Mutant_3 showed maximum H-bonding and better stability than other mutants and WT that proved higher affinity suggesting improved catalysis. Mutant_2 and Mutant_3 exhibited binding affinities of −7.6 and −8.2 kcal/mol, respectively for the ligand. The outcome of present study will provide significant improvement in synthesis of kaempferol and other plant-based flavonoids.

Communicated by Ramaswamy H. Sarma

Keywords:

• MD simulation
• molecular docking
• PCA
• DCCM
• FEL
• MM_PBSA

Acknowledgement

The facility provided by DBT, GOI, through BTISnet SubDIC to Department of the Bioengineering and Biotechnology, Birla Institute of Technology, Mesra, Ranchi, India for molecular dynamics studies is highly acknowledged.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

DST/INSPIRE Fellowship (as Junior Research Fellowship) awarded to GK by Government of India, Ministry of Science and Technology, Department of Science and Technology Bhawan, New Mehrauli Road, New Delhi-110016, India having Grant No [IF180386] supports financially.