Structure-function analysis of a series of novel GIP analogues containing different helical length linkers.
Glucose-dependent insulinotropic polypeptide (GIP1-42) is a potent glucose-lowering intestinal peptide hormone. The equipotent GIP1-30NH2 was structurally modified by linking N- and C-terminal fragments with several different linkers. Substitution of the middle region of GIP by a flexible aminohexanoic linker resulted in greatly reduced binding affinity and reduction or complete loss of bioactivity. Connection of the bioactive domains GIP1-14 and GIP19-30NH2 by EKEK or AAAA linkers resulted in peptide agonists with approximately 3-4-fold increased bioactivity as compared to GIP1-30NH2. Conformational analysis by CD spectroscopy of GIP fragments and analogues suggests a helical region in the C-terminal (19-30) portion of GIP. It was demonstrated that stabilization of this C-terminal helical region by the introduction of helical linkers favored binding and activation of the GIP receptor. Our results suggest an important contribution of a direct interaction of the first 14 amino acids with the GIP receptor, an appropriate relative orientation of N- and C-terminal parts of GIP, and the presence of helical linkers to be essential for bioactivity.
Manhart
Susanne
S
Department of Peptide Chemistry, probiodrug AG, Weinbergweg 22, 06120 Halle/Saale, Germany.
Hinke
Simon A
SA
McIntosh
Christopher H S
CH
Pederson
Raymond A
RA
Demuth
Hans-Ulrich
HU
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D016466:CHO Cells; D002942:Circular Dichroism; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism; D005749:Gastric Inhibitory Polypeptide / Q000138:chemical synthesis* / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000502:physiology; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011869:Radioligand Assay; D051381:Rats; D013329:Structure-Activity Relationship; D014162:Transfection
D010446:Peptide Fragments; D005749:Gastric Inhibitory Polypeptide; D000242:Cyclic AMP