Rat guanylyl cyclase C expressed in COS-7 cells exhibits multiple affinities for Escherichia coli heat-stable enterotoxin.
Rat guanylyl cyclase C expressed in COS-7 cells exhibits multiple affinities for Escherichia coli heat-stable enterotoxin.
复制标题
在 COS-7 细胞中表达的大鼠鸟苷酸环化酶 C 对大肠杆菌热稳定肠毒素表现出多种亲和力。
DOI：
10.1021/bi00028a019
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发表时间：
1995
期刊：
影响因子：
2.9
通讯作者：
Waldman,SA
中科院分区：
文献类型：
--
作者：
Deshmane,SP;Carrithers,SL;Parkinson,SJ;Crupper,SS;Robertson,DC;Waldman,SA
Revised Manuscript Received May 15, 1995® abstract: Intestinal cells exhibit binding sites with different affinities for Escherichia coli heat-stable enterotoxin (ST) and guanylin, suggesting the existence of different receptors for these peptides. Guanylyl cyclase C from intestinal cells has been identified as one receptor for these peptides. Equilibrium and kinetic binding characteristics of rat guanylyl cyclase C expressed in COS-7cells were examined, employing ST, to determine if this receptor exhibited multiple affinities. Scatchard analysis of equilibrium binding yielded curvilinear isotherms consistent with the presence of high (pM) and low (nM) affinity sites. Kinetic analysis of binding demonstrated that these sites exhibitedsimilar dissociation but different association kinetics. In addition, two distinct affinity states of low affinity sites were identified with dissociation constants of 0.15 and 5.85 nM. Association of ST and low affinity sites was biphasic, while dissociation from these sites was unimodal. Close agreement of equilibrium and kinetic dissociationconstants suggested that low affinity sites were in the lowest affinity state at equilibrium. Comparison of the ligand dependence of guanylyl cyclase activity (EC50= 110 nM) with receptor occupancy revealed that binding of ST to the lowest affinity state of low affinity sites (EC50= 80 nM) is directly coupled to catalytic activation. These studies suggest that binding sites with different affinities for ST exhibited by intestinal cells reflect the expression of a single gene product, guanylyl cyclase C, rather than different receptors for the ligand. The shiftin affinity state of low affinity sites and its correlation with catalytic activation suggest a central role for this phenomenon in mechanisms mediating receptor—effector coupling of membrane guanylyl cyclases.
