Role of Cel5H protein surface amino acids in binding with clay minerals and measurements of its forces

• Published in: Applied microscopy Vol. 51; no. 1; p. 17
• Main Authors: Math, Renukaradhya K., Bharatham, Nagakumar, Javaregowda, Palaksha K., Yun, Han Dae
• Format: Journal Article
• Language: English
• Published: Singapore Springer Singapore 11-11-2021; Springer Nature B.V; SpringerOpen; 한국현미경학회
• Subjects: Adhesion force; AFM; Amino acids; Atomic force microscopy; Binding; Cellulase; Chemistry and Materials Science; Clay; Clay mineral; Clay minerals; Electrostatic properties; Force measurement; Homology modeling; Lysine; Materials Science; Mica; Minerals; Mutants; Mutation; Protein binding; Proteins; Residues; 자연과학일반; Adhesion force; AFM; Mutation; Clay mineral; Homology modeling; Protein binding
• ISSN: 2287-4445; 2287-5123; 2287-4445
• DOI: 10.1186/s42649-021-00066-7

Our previous study on the binding activity between Cel5H and clay minerals showed highest binding efficiency among other cellulase enzymes cloned. Here, based on previous studies, we hypothesized that the positive amino acids on the surface of Cel5H protein may play an important role in binding to clay surfaces. To examine this, protein sequences of
Bacillus licheniformis
Cel5H (
Bl
Cel5H) and
Paenibacillus polymyxa
Cel5A (
Pp
Cel5A) were analyzed and then selected amino acids were mutated. These mutated proteins were investigated for binding activity and force measurement via atomic force microscopy (AFM). A total of seven amino acids which are only present in
Bl
Cel5H but not in
Pp
Cel5A were selected for mutational studies and the positive residues which are present in both were omitted. Of the seven selected surface lysine residues, only three mutants K196A(M2), K54A(M3) and K157T(M4) showed 12%, 7% and 8% less clay mineral binding ability, respectively compared with wild-type. The probable reason why other mutants did not show altered binding efficiency might be due to relative location of amino acids on the protein surface. Meanwhile, measurement of adhesion forces on mica sheets showed a well-defined maximum at 69 ± 19 pN for wild-type, 58 ± 19 pN for M2, 53 ± 19 pN for M3, and 49 ± 19 pN for M4 proteins. Hence, our results demonstrated that relative location of surface amino acids of Cel5H protein especially positive charged amino acids are important in the process of clay mineral-protein binding interaction through electrostatic exchange of charges.