Crystallization and preliminary X-ray analysis of the complexes between a Fab and two forms of human insulin-like growth factor II.
Elevated expression of insulin-like growth factor II (IGF-II) is frequently observed in a variety of human malignancies, including breast, colon and liver cancer. As IGF-II can deliver a mitogenic signal through both the type 1 insulin-like growth factor receptor (IGF-IR) and an alternately spliced form of the insulin receptor (IR-A), neutralizing the biological activity of this growth factor directly is an attractive therapeutic option. One method of doing this would be to find antibodies that bind tightly and specifically to the peptide, which could be used as protein therapeutics to lower the peptide levels in vivo and/or to block the peptide from binding to the IGF-IR or IR-A. To address this, Fabs were selected from a phage-display library using a biotinylated precursor form of the growth factor known as IGF-IIE as a target. Fabs were isolated that were specific for the E-domain C-terminal extension and for mature IGF-II. Four Fabs selected from the library were produced, complexed with IGF-II and set up in crystallization trials. One of the Fab-IGF-II complexes (M64-F02-IGF-II) crystallized readily, yielding crystals that diffracted to 2.2 A resolution and belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 50.7, b = 106.9, c = 110.7 A. There was one molecule of the complete complex in the asymmetric unit. The same Fab was also crystallized with a longer form of the growth factor, IGF-IIE. This complex crystallized in space group P2(1)2(1)2(1), with unit-cell parameters a = 50.7, b = 107, c = 111.5 A, and also diffracted X-rays to 2.2 A resolution.
Newman
Janet
J
Molecular and Health Technologies, CSIRO, Parkville, Victoria, Australia. janet.newman@csiro.au
Cohen
Edward H
EH
Cosgrove
Leah
L
Kopacz
Kris
K
Dransfield
Daniel T
DT
Adams
Timothy E
TE
Peat
Thomas S
TS
D003460:Crystallization; D018360:Crystallography, X-Ray; D006031:Glycosylation; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D007335:Insulin-Like Growth Factor II / Q000737:chemistry*; D020033:Protein Isoforms / Q000737:chemistry
D007140:Immunoglobulin Fab Fragments; D020033:Protein Isoforms; D007335:Insulin-Like Growth Factor II