We have identified structural attributes required for signal transduction through a seven-transmembrane-domain receptor. Platelets from a patient (AC) with a congenital bleeding disorder had normal shape change but reduced and reversible aggregation in response to 4 μM ADP, similar to normal platelets with blocked P2Y₁₂ receptor. The response to 20 μM ADP, albeit still decreased, was more pronounced and was reduced by a P2Y₁₂ antagonist, indicating some residual receptor function. ADP failed to lower the adenylyl cyclase activity stimulated by prostaglandin E₁ in the patient's platelets, even though the number and affinity of 2-methylthioadenosine 5′-[³³P]diphosphate-binding sites was normal. Analysis of the patient's P2Y₁₂ gene revealed a G-to-A transition in one allele, changing the codon for Arg-256 in the sixth transmembrane domain to Gln, and a C-to-T transition in the other allele, changing the codon for Arg-265 in the third extracellular loop to Trp. Neither mutation interfered with receptor surface expression but both altered function, since ADP inhibited the forskolin-induced increase of cAMP markedly less in cells transfected with either mutant P2Y₁₂ as compared with wild-type receptor. These studies delineate a region of P2Y₁₂ required for normal function after ADP binding.