An unusual cysteine V L 87 affects the antibody fragment conformations without interfering with the disulfide bond formation

ATTALLAH, CAROLINA; AGUILAR, MARÍA FERNANDA; GARAY, A. SERGIO; HERRERA, FERNANDO E.; ETCHEVERRIGARAY, MARINA; OGGERO, MARCOS; RODRIGUES, DANIEL E.

MOLECULAR IMMUNOLOGY

PERGAMON-ELSEVIER SCIENCE LTD

Año: 2017 vol. 90 p. 143 - 149

0161-5890

p { margin-bottom: 2.47mm; direction: ltr; color: rgb(0, 0, 10); line-height: 120%; widows: 2; orphans: 2; }p.western { font-family: "Calibri",sans-serif; font-size: 11pt; }p.cjk { font-family: "Calibri",sans-serif; font-size: 11pt; }p.ctl { font-family: "Arial",sans-serif; font-size: 11pt; }a:link { color: rgb(5, 99, 193); }a.western:link { }a.ctl:link { }TheCys residues are almost perfectly conserved in all antibodies. Theycontribute significantly to the antibody fragment stability. Therelevance of two natural contiguous Cys residues of ananti‑recombinant human‑follicle stimulation hormone(rhFSH) in a format of single‑chain variable fragment (scFv)was studied. This scFv contains 5 Cys residues: VH22and VH92in the variable heavy chain (VH)and VL23,VL87and VL88in the variable light chain (VL).The influence of two unusual contiguous Cys at positions VL87and VL88was studied by considering the wild type fragment and mutantvariants: VL‑C88S,VL‑C87S,VL‑C87Y.The analysis was carried out using antigen‑binding abilitymeasurement by indirect specific ELISA and a detailed molecularmodeling that comprises homology methods, long molecular dynamicssimulations and docking. We found that VL‑C87affected the antibody fragment stability without interfering with thedisulfide bond formation.Theeffect of mutating the VL‑C87by a usual residue at this position like Tyr caused distantstructural changes at the VHregion that confers a higher mobility to the VH‑CDR2and VH‑CDR3loops improving the scFv binding to the antigen.