James F. Perdue

DEVELOPMENT OF A SPECIFIC RADIOIMMUNO ASSAY FOR E DOMAIN CONTAINING FORMS OF INSULIN-LIKE GROWTH FACTOR II

The analysis of cDNA clones for human (1,2) and rat (3) insulin-like growth factor-II (IGF-II)1 has led to the prediction that the processed forms of the growth factors, i.e. Mr = 7422 (67 amino acids) for human IGF-II are synthesized as precursors with an extension of 89 amino acids at the carboxyl terminus. This extension is termed the E domain. Moses et al (4) identified two precursor forms of rat IGF-II (originally designated multiplication-stimulating activity) in the conditioned medium of Buffalo rat liver, i.e. BRL-3A, cells with appMrs = 16,270 (MSA-1) and 8,700 (MSA-II). Human serum, spinal fluid and tissue extracts also contain high Mr forms of IGF-II (5-8). Zumstein et al., (5) have purified a Mr = 10,000 variant form of IGF-II from serum that contained Cys-Gly-Asp for Ser33 in the C domain and an E domain extension of 21- amino acids. This 10 kDa IGF-II may be similar or identical to the “big IGF-II” that was reported to be present in human serum and in spinal fluid (6). We have isolated a still larger form of IGF-II from normal human serum (7). N-terminal amino acid sequence analysis through the first 28 residues and RRAs using rat placental membranes established it as a form of IGF-II. As evidenced from its mobility during SDS-PAGE, it has an appMr = 15,000. Very recently, Hudgins et al., (8) established that normal human serum contains several forms of precursor IGF- II with acidic isoelectric points, i.e. pI’s. The mass and acidic nature of one of these molecules with an apparent Mr= 15,000 was contributed, in part, by polysaccharides and sialic acids, respectively (8). These results may explain the observations of several investigators that extracts from the tissues and serum of patients with malignant tumors contain a broad size range of IGF-II (2,9,10).

Towards Identification of a Binding Site on Insulin-Like Growth Factor-II for IGF-Binding Proteins

Insulin-like growth factor (IGF) -I and -II mediate mitogenesis, differentiation and insulin-like metabolic effects.1 Their amino acid sequence is divided into 4 domains designated B-C-A-D beginning from the amino terminus. The B-and A-domains of IGF-I and IGF-II share substantial homology with each other and with the B-and A-chains of insulin.