Crystallization and structure determination of an autoimmune antipoly(dT) immunoglobulin Fab fragment at 3.0 Å resolution

M. Cygler, A. Boodhoo, J. S. Lee, W. F. Anderson

Research output: Contribution to journalArticlepeer-review

73 Scopus citations

Abstract

HED10 is an autoimmune antibody (IgG) which shows considerable specificity for the single-stranded DNA poly(dT). Production of Fab fragments by papain digestion resulted in heterogeneity as judged by isoelectric focusing gels, which had a marked negative effect on crystallization. However, a single species of Fab with a pI of 7.6 could be isolated in good yield by DEAE-cellulose chromatography, and good crystals were produced by the hanging drop vapor diffusion method. The space group was P21 with cell dimensions, a = 64.2, b = 90.0, c = 42.3 Å, and β = 96.7°. These crystals diffract to about 2.2 Å resolution. The structure of Fab HED10 was solved by the molecular replacement method using the known structure of McPC603 and is refined to R = 27.2% at 3.0 Å resolution. Fab HED10 is more extended than McPC603 and has an elbow angle (between the variable and constant domains) of 162° , very similar to that observed in Fab KOL. The majority of the hypervariable regions are visible in the model.

Original languageEnglish (US)
Pages (from-to)643-648
Number of pages6
JournalJournal of Biological Chemistry
Volume262
Issue number2
StatePublished - Mar 26 1987

ASJC Scopus subject areas

  • Biochemistry

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