Crystal structure of the disulfide-stabilized Fv fragment of anticancer antibody B1: Conformational influence of an engineered disulfide bond

Orna Almog, Itai Benhar, George Vasmatzis, Maria Tordova, Byungkook Lee, Ira Pastan, Gary L. Gilliland

Research output: Contribution to journalArticlepeer-review

16 Scopus citations


A recombinant Fv construct of the B1 monoclonal antibody that recognizes the Lewis(Y)-related carbohydrate epitope on human carcinoma cells has been prepared. The Fv is composed of the polypeptide chains of the V(H) and V(L) domains expressed independently and isolated as inclusion bodies. The Fv is prepared by combining and refolding equimolar amounts of guanidine chloride solubilized inclusion bodies. The Fv is stabilized by an engineered interchain disulfide bridge between residues V(L)100 and V(H)44. This construct has a similar binding affinity as that of the single-chain construct (Benhar and Pastan, Clin. Cancer Res. 1:1023-1029, 1995). The B1 disulfide-stabilized Fv (BldsFv) crystallizes in space group P6122 with the unit cell parameters a = b = 80.1 Å, and c = 138.1 Å. The crystal structure of the BldsFv has been determined at 2.1-Å resolution using the molecular replacement technique. The final structure has a crystallographic R-value of 0.187 with a root mean square deviation in bond distance of 0.014 and in bond angle of 2.74°. Comparisons of the BldsFv structure with known structures of Fv regions of other immunoglobulin fragments shows closely related secondary and tertiary structures. The antigen combining site of BldsFv is a deep depression 10-Å wide and 17-Å long with the walls of the depression composed of residues, many of which are tyrosines, from complementarity determining regions L1, L3, H1, H2, and H3. Model building studies indicate that the Lewis(Y) tetrasaccharide, Fuc-Gal-Nag-Fuc, can be accommodated in the antigen combining site in a manner consistent with the epitope predicted in earlier biochemical studies (Pastan, Lovelace, Gallo, Rutherford, Magnani, and Willingham, Cancer Res. 51:3781-3787, 1991). Thus, the engineered disulfide bridge appears to cause little, if any, distortion in the Fv structure, making it an effective substitute for the B1 Fab.

Original languageEnglish
Pages (from-to)128-138
Number of pages11
JournalProteins: Structure, Function and Bioinformatics
Issue number2
StatePublished - 1 May 1998
Externally publishedYes


  • Antibody
  • Antitumor
  • Protein stability
  • Protein structure
  • Single chain Fv
  • Variable domains
  • X-ray crystallography

ASJC Scopus subject areas

  • Structural Biology
  • Biochemistry
  • Molecular Biology


Dive into the research topics of 'Crystal structure of the disulfide-stabilized Fv fragment of anticancer antibody B1: Conformational influence of an engineered disulfide bond'. Together they form a unique fingerprint.

Cite this