A molecular dynamics study of ligand binding in mutant avidins
KUKKURAINEN, SALLI (2009)
2009
Bioinformatiikka - Bioinformatics
Lääketieteellinen tiedekunta - Faculty of Medicine
This publication is copyrighted. You may download, display and print it for Your own personal use. Commercial use is prohibited.
Hyväksymispäivämäärä
2009-03-02
Julkaisun pysyvä osoite on
https://urn.fi/urn:nbn:fi:uta-1-19620
https://urn.fi/urn:nbn:fi:uta-1-19620
Tiivistelmä
Background and aims: Chicken avidin is a homotetrameric β-barrel protein, whose affinity towards its natural ligand biotin is the strongest non-covalent protein-ligand interaction known. Functional avidin has four identical cavities that can bind small, hydrophobic molecules. Residues lining these cavities have been selected as targets for mutagenesis in studies aiming towards the development of novel avidin-based protein structures with different functions. Previous studies using site-directed mutagenesis of loop regions have yielded testosterone-binding avidin structures. In addition, two point mutations have been found to markedly reduce the affinity of avidin towards biotin. In this study, I have used computational methods to model the molecular interactions of two mutant avidins named NRMNH and NQNGY, with testosterone, and those of two keto-biotin binders, T35P,N118M and T35K,N118M, with biotin and keto-biotin.
Methods: A known 3-D sturucture of avidin was used as a template in the modeling of mutant avidin structures. The structural models of biotin, keto-biotin, and testosterone were placed in the ligand-binding pocket of the protein structures by molecular docking. Molecular interactions in the protein-ligand complexes were analyzed by molecular dynamics simulations. Hydrogen bonding patterns, ligand motion, the mobilities of individual residues as a function of time and with respect to those of apo-avidin were calculated, and protein-ligand contact area was estimated.
Results: Two candidate binding modes for testosterone binding in avidin-based structures were identified using calculations of ligand and residue motion and hydrogen bonding patterns. Both of these modes were observed in the NRMNH mutant, one in NQNGY, and one in wt-Avd. Studies on keto-biotin binding structures showed that the T35P,N118M mutant structure may bind biotin weaker than wt-Avd or the N118M mutant. Keto-biotin and biotin were not found to bind to T35K,N118M.
Conclusions: Molecular dynamics simulations were found to be a potential tool for studying ligand binding in avidin-based structures. Hydrogen bonding patterns, ligand motion, and protein residue motion were found as applicable analysis methods. Individual amino acids were pointed out for further mutagenesis, to yield better binders for testosterone. The results gained should be confirmed computationally and experimentally.
Methods: A known 3-D sturucture of avidin was used as a template in the modeling of mutant avidin structures. The structural models of biotin, keto-biotin, and testosterone were placed in the ligand-binding pocket of the protein structures by molecular docking. Molecular interactions in the protein-ligand complexes were analyzed by molecular dynamics simulations. Hydrogen bonding patterns, ligand motion, the mobilities of individual residues as a function of time and with respect to those of apo-avidin were calculated, and protein-ligand contact area was estimated.
Results: Two candidate binding modes for testosterone binding in avidin-based structures were identified using calculations of ligand and residue motion and hydrogen bonding patterns. Both of these modes were observed in the NRMNH mutant, one in NQNGY, and one in wt-Avd. Studies on keto-biotin binding structures showed that the T35P,N118M mutant structure may bind biotin weaker than wt-Avd or the N118M mutant. Keto-biotin and biotin were not found to bind to T35K,N118M.
Conclusions: Molecular dynamics simulations were found to be a potential tool for studying ligand binding in avidin-based structures. Hydrogen bonding patterns, ligand motion, and protein residue motion were found as applicable analysis methods. Individual amino acids were pointed out for further mutagenesis, to yield better binders for testosterone. The results gained should be confirmed computationally and experimentally.