Isolation and characterization of circulating 13-kDa C-terminal fragments of human insulin-like growth factor binding protein-5.
The insulin-like growth factor binding proteins (IGFBPs) are responsible for regulation of the effects and the bioavailability of the insulin-like growth factors (IGFs). We screened for circulating fragments of human IGFBP-5 in human hemofiltrate. Identification of IGFBP-5 peptides in the fractions of our peptide bank generated from hemofiltrate was performed by their immunoreactivity and their capacity to bind IGF-I. Different fragments of IGFBP-5 with molecular sizes from 12 to 25 kDa were identified. C-terminal peptides of IGFBP-5 with molecular masses of 13.3 and 13.5 kDa were purified by consecutive chromatographic steps and sequenced. Sequence analysis of the peptides revealed the (double) sequences (K)FVGGAENXAHPRII and MVPRAVYLPNXDRKG. In addition, a smaller fragment with Mr 2722 of the central IGFBP-5 region was purified and showed the sequence HTRISELKAEAVKKDRRKKLTQS (residues 121-143) indicating plasma proteolysis of IGFBP-5 C-terminal to amino acids Lys-120, Ser-143, Lys-144, and Arg-188. According to mass spectrometric and sequence analysis, Thr-152 was shown to be O-glycosylated. Fractions containing C-terminal IGFBP-5 fragments revealed significant IGF-I binding properties. Our results indicate that plasma proteolysis of IGFBP-5 preferentially occurs C-terminally to basic residues and generates different C-terminal fragments, possibly acting in an IGF-dependent manner and bearing intrinsic biological functions.
Ständker
L
L
Lower Saxony Institute for Peptide Research, Hannover, Germany. 106535.324@compuserve.com
Wobst
P
P
Mark
S
S
Forssmann
W G
WG
D000595:Amino Acid Sequence; D006031:Glycosylation; D006801:Humans; D006868:Hydrolysis; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000097:blood / Q000737:chemistry*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000097:blood / Q000737:chemistry / Q000302:isolation & purification*
D018975:Insulin-Like Growth Factor Binding Protein 5; D010446:Peptide Fragments