Promoting crystallization of antibody-antigen complexes via microseed matrix screening.
The application of microseed matrix screening to the crystallization of antibody-antigen complexes is described for a set of antibodies that include mouse anti-IL-13 antibody C836, its humanized version H2L6 and an affinity-matured variant of H2L6, M1295. The Fab fragments of these antibodies were crystallized in complex with the antigen human IL-13. The initial crystallization screening for each of the three complexes included 192 conditions. Only one hit was observed for H2L6 and none were observed for the other two complexes. Matrix self-microseeding using these microcrystals yielded multiple hits under various conditions that were further optimized to grow diffraction-quality H2L6 crystals. The same H2L6 seeds were also successfully used to promote crystallization of the other two complexes. The M1295 crystals appeared to be isomorphous to those of H2L6, whereas the C836 crystals were in a different crystal form. These results are consistent with the concept that the conditions that are best for crystal growth may be different from those that favor nucleation. Microseed matrix screening using either a self-seeding or cross-seeding approach proved to be a fast, robust and reliable method not only for the refinement of crystallization conditions but also to promote crystal nucleation and increase the hit rate.
Obmolova
Galina
G
Centocor R&D, 145 King of Prussia Road, Radnor, PA 19087, USA. gobmolov@its.jnj.com
Malia
Thomas J
TJ
Teplyakov
Alexey
A
Sweet
Raymond
R
Gilliland
Gary L
GL
D000818:Animals; D000936:Antigen-Antibody Complex / Q000737:chemistry* / Q000276:immunology; D003460:Crystallization; D006801:Humans; D018793:Interleukin-13 / Q000737:chemistry* / Q000276:immunology; D051379:Mice
D000936:Antigen-Antibody Complex; D018793:Interleukin-13