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N-terminal deletion mutants of insulin-like growth factor-II (IGF-II) show Thr7 and Leu8 important for binding to insulin and IGF-I receptors and Leu8 critical for all IGF-II functions. 1995

R Hashimoto, and H Fujiwara, and N Higashihashi, and T Enjoh-Kimura, and H Terasawa, and Y Fujita-Yamaguchi, and F Inagaki, and J F Perdue, and K Sakano
Molecular Biology Research Laboratory, Daiichi Pharmaceutical Co., Ltd., Tokyo, Japan.

To define the role of the N-terminal region of insulin-like growth factor-II (IGF-II) in its binding to insulin and IGF receptors, deletion mutants des-(1-5)-, des-(1-7)-, and des-(1-8)-recombinant (r) IGF-II, and the Gly8 for Leu substitution mutant of rIGF-II were prepared by site-directed mutagenesis, expressed in Escherichia coli, and purified. The binding affinity and mitogenic activity of these rIGF-II mutants as well as commercially available des-(1-6)-rIGF-II were analyzed. While the relative affinity of des-(1-5)- and des-(1-6)-rIGF-II for purified human insulin and IGF-I receptors remained at > or = 50% levels of that of rIGF-II, the affinity of des-(1-7)-rIGF-II decreased to approximately 10% and approximately 3%, respectively, of that of rIGF-II. When the octapeptide including Leu8 was removed prior to the Cys9-Cys47 intrachain bond, the relative affinity of this deletion mutant, des-(1-8)-rIGF-II, for these receptors dramatically decreased to < 1% of that of rIGF-II. Substituting Gly8 for Leu in rIGF-II decreased the affinity of this mutant for the IGF-I and insulin receptors to about the same extent. These results suggest that the side chains of Thr7 and Leu8 may play an important role in retaining all of the IGF-II functions. Decreases in the relative affinity for binding of the mutants to these receptors paralleled the decreases in their mitogenic potency for cultured Balb/c 3T3 cells. Although the relative affinity of des-(1-8)- or [Gly8]rIGF-II for rat IGF-II/CIM6-P (cation-independent mannose 6-phosphate) receptors was also < 1% of that of rIGF-II, the relative affinities of des-(1-5)-, des-(1-6)-, and des-(1-7)-rIGF-II for these receptors was significantly greater than that of rIGF-II. These results clearly demonstrate that Thr7 and Leu8 are important for binding to insulin and IGF-I receptors and Leu8 is critical for expression of all IGF-II functions.

UI MeSH Term Description Entries
D007335 Insulin-Like Growth Factor II A well-characterized neutral peptide believed to be secreted by the LIVER and to circulate in the BLOOD. It has growth-regulating, insulin-like and mitogenic activities. The growth factor has a major, but not absolute, dependence on SOMATOTROPIN. It is believed to be a major fetal growth factor in contrast to INSULIN-LIKE GROWTH FACTOR I, which is a major growth factor in adults. IGF-II,Multiplication-Stimulating Activity,Somatomedin MSA,IGF-2,Insulin Like Growth Factor II,Insulin-Like Somatomedin Peptide II,Multiplication-Stimulating Factor,Somatomedin A,Factor, Multiplication-Stimulating,Insulin Like Somatomedin Peptide II,Multiplication Stimulating Activity,Multiplication Stimulating Factor
D007930 Leucine An essential branched-chain amino acid important for hemoglobin formation. L-Leucine,Leucine, L-Isomer,L-Isomer Leucine,Leucine, L Isomer
D008807 Mice, Inbred BALB C An inbred strain of mouse that is widely used in IMMUNOLOGY studies and cancer research. BALB C Mice, Inbred,BALB C Mouse, Inbred,Inbred BALB C Mice,Inbred BALB C Mouse,Mice, BALB C,Mouse, BALB C,Mouse, Inbred BALB C,BALB C Mice,BALB C Mouse
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D011485 Protein Binding The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments. Plasma Protein Binding Capacity,Binding, Protein
D011972 Receptor, Insulin A cell surface receptor for INSULIN. It comprises a tetramer of two alpha and two beta subunits which are derived from cleavage of a single precursor protein. The receptor contains an intrinsic TYROSINE KINASE domain that is located within the beta subunit. Activation of the receptor by INSULIN results in numerous metabolic changes including increased uptake of GLUCOSE into the liver, muscle, and ADIPOSE TISSUE. Insulin Receptor,Insulin Receptor Protein-Tyrosine Kinase,Insulin Receptor alpha Subunit,Insulin Receptor beta Subunit,Insulin Receptor alpha Chain,Insulin Receptor beta Chain,Insulin-Dependent Tyrosine Protein Kinase,Receptors, Insulin,Insulin Receptor Protein Tyrosine Kinase,Insulin Receptors
D004247 DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA, Double-Stranded,Deoxyribonucleic Acid,ds-DNA,DNA, Double Stranded,Double-Stranded DNA,ds DNA
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000595 Amino Acid Sequence The order of amino acids as they occur in a polypeptide chain. This is referred to as the primary structure of proteins. It is of fundamental importance in determining PROTEIN CONFORMATION. Protein Structure, Primary,Amino Acid Sequences,Sequence, Amino Acid,Sequences, Amino Acid,Primary Protein Structure,Primary Protein Structures,Protein Structures, Primary,Structure, Primary Protein,Structures, Primary Protein
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia

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