Molecular modeling and docking analysis of the variable regions of an anti-N6-methyladenosine monoclonal antibody.

摘要

The previously unknown DNA and amino acid sequences of the 7C7:C5 anti-N 6-methyladenosine (6mAos) variable fragment (Fv) are reported herein. The 7C7:C5 Fv was molecular modeled using a novel approach involving alignment of numerous high identity antigen-bound crystallographic structures and antigen-unbound crystallographic structures as templates, in which a binding pocket was revealed only in the antigen-bound-model of the Fv. Rigid Fv/flexible antigen docking and flexible Fv/flexible antigen docking of deoxy-N6-methyladenosine (d6mA), deoxyadenosine (dA), and single-stranded (ss) DNA trinucleotide (A- 6mA-T) revealed three critical residues (TYR L48, SER L55, and PRO H103) to be involved in docking of d6mA and related antigens to the 7C7:C5 Fv. The N6 methyl group in d6mA was predicted to make more van der Waals contacts than the corresponding N6 hydrogen in dA leading to an estimated 6 to 10 fold greater affinity for d6mA than dA. Intermediate dissociation constant (Kd) calculations suggest a Kd range of 7.835 x 10-7 M to 5.92 x 10 -8 M for the binding of d6mA in ssDNA, which corresponds to an affinity most appropriately used for genotyping or diagnostics. The binding interactions suggest the possibility of a slightly higher affinity for ribonucleoside versus deoxyribonucleoside, suggesting that the 7C7:C5 Fv may be able to bind to N6-methyladenosine (6mA os) in RNA.