To determine which residues within the rat somatostatin receptor subtype SSTR2 may be interacting with the lys9 of somatostatin-14 (S-14), mutant SSTR2 receptors were created by mutating asp89 or asp122. [125I Tyr11]S-14 binding to D89A and D89E mutants suggests that asp89 is not directly involved in S-14 binding. Binding studies with the charge switch mutants, asp9S-14, and D122K, suggest that asp122 may be interacting with the lys9 of S-14. [125I Tyr11]asp9S-14 displayed saturable binding to D122K with an affinity comparable to that seen with [125I Tyr11]S-14 and WT SSTR2. These data suggest that the interaction between lys9 of S-14 and the TM3 asp122 of SSTR2 represents one contact site between S-14 and SSTR2.