Structurefunction Analysis Of sstr2

Presently there are major limitations of SRIF receptor pharmacology. The lack of antagonists specific for each receptor subtype has greatly limited studies designed to identify the selective functions of SRIF receptors. Furthermore, no nonpeptide SRIF ligands are commonly available. As a result, SRIF analogs as therapeutic agents is limited. Conceivably, with the availability of the cloned receptors, pharmaceutical companies may advance in the development of these agents.

Structure-function analysis of the ligand binding domains of the receptors may aid in development of new SRIF ligands. Recently, several studies have focussed on ligand binding determinants of sstr2. Fitzpatrick and Vandlen (43) showed that the second and third extracellular loops of sstr2 are critical for the binding of MK 678 and Kaupmann et al. (44) reported that several amino acids in the vicinity of these loops were essential for the binding of octreotide to the receptor. Liapakis et al. (45) reported that a four amino-acid sequence phenylalanine-aspartate-phenylalanine-valine (FDFV) at the interphase of the third extracellular loop and transmembrane seven of sstr2 was involved in the binding of octapeptide and hexapeptides to sstr2. A phenylalanine at residue 294 was most essential for the binding of octapeptides, because this residue inserted into a corresponding region of sstr1 to create the mutant sstr1S305F conferred onto sstr1 the ability to bind octapeptides.

Hexapeptides, such as MK 678, did not bind to sstr1 S305F, suggesting that this smaller SRIF analog had different requirements for binding than octapeptides. MK 678 has a tyrosine adjacent to the tryptophan and lysine residues needed for binding. Hexapeptide analogs with a phenylalanine at this position bound to sstr1S305F, indicating that the phenylalanine of the peptide was critical for the ability of the peptide to interact with the phenylalanine at residue 294 of sstr1. Conceivably, the phenylalanine allows for hydrophobic interactions to occur with the phenylalanine of the receptor, whereas the added hydroxyl group of MK 678s tyrosine serves to repel the peptide from the phenylalanine of the receptor. This structural information reveals both critical determinants of the peptides and receptor needed for binding, which could be useful in the design of new sstr2 select ligands.

The mutagenesis studies also revealed determinants for binding to sstr1 (45). The peptide des-AA125-[DTrp8,IAMP9]SRIF binds selectively to sstr1 and does not bind to sstr2. The peptide bound to a chimeric receptor consisting of sstr2 with the second extracellular loop of sstr1, suggesting that this region of sstr1 is essential for the binding of this synthetic analog. Thus, the binding domains of selective agonists at sstr1 and sstr2 are at vastly different regions of the receptor.

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