Fonteles M C, Carrithers S L, Monteiro H S, Carvalho A F, Coelho G R, Greenberg R N, Forte L R
Clinical Research Unit of Federal University of Ceara and Ceara State University, 60434 Fortaleza-CE, Brazil.
Am J Physiol Renal Physiol. 2001 Feb;280(2):F207-13. doi: 10.1152/ajprenal.2001.280.2.F207.
Guanylin and uroguanylin compose a family of natriuretic, diuretic, and kaliuretic peptides that bind to and activate apical membrane receptor guanylyl cyclase signaling molecules in renal and intestinal epithelia. Recently, a complementary DNA encoding an additional member of the guanylin family of cGMP-regulating peptides was isolated from lymphoid tissues of the opossum and was termed lymphoguanylin (LGN). A peptide analog of opossum LGN was synthesized containing a single disulfide bond with the internal cysteine-7 replaced by a serine residue (LGN(Cys7-->Ser7)). The biological activity of LGN(Ser) was tested by using a cGMP bioassay with cultured T84 (human intestinal) cells and opossum kidney (OK) cells. LGN(Ser) has potencies and efficacies for activation of cGMP production in the intestinal and kidney cell lines that are 100- and 1,000-fold higher than LGN, respectively. In the isolated perfused rat kidney, LGN(Ser) stimulated a maximal increase in fractional Na+ excretion from 24.8 +/- 3.0 to 36.3 +/- 3.3% 60 min after administration and enhanced urine flow from 0.15 +/- 0.01 to 0.24 +/- 0.01 ml. g(-1). min(-1). LGN(Ser) (0.69 microM) also increased fractional K+ excretion from 27.3 +/- 2.3 to 38.0 +/- 3.0% and fractional Cl- excretion from 26.1 +/- 0.8 to 43.5 +/- 1.9. A ninefold increase in the urinary excretion of cGMP from 1.00 +/- 0.04 to 9.28 +/- 1.14 pmol/ml was elicited by LGN(Ser), whereas cAMP levels were not changed on peptide administration. These findings demonstrate that LGN(Ser), which contains a single disulfide bond like native LGN, activates guanylyl cyclase-C (GC-C) receptors in T84 and OK cells and may be very helpful in studying the physiological importance of activation of GC-C in vivo. LGN(Ser) also exhibits full activity in the isolated perfused kidney equivalent to that observed previously with opossum uroguanylin, suggesting a physiological role for LGN in renal function. Thus the single amino acid substitution enhances the activity and potency of LGN.
鸟苷林和尿鸟苷林构成了一类利钠、利尿和利钾肽家族，它们能与肾和肠上皮细胞的顶端膜受体鸟苷酸环化酶信号分子结合并激活这些分子。最近，从负鼠的淋巴组织中分离出一种编码cGMP调节肽鸟苷林家族另一个成员的互补DNA，它被命名为淋巴鸟苷林（LGN）。合成了一种负鼠LGN的肽类似物，其含有一个单一的二硫键，内部的半胱氨酸 - 7被丝氨酸残基取代（LGN(Cys7→Ser7)）。通过使用培养的T84（人肠）细胞和负鼠肾（OK）细胞的cGMP生物测定法来测试LGN(Ser)的生物活性。LGN(Ser)在肠和肾细胞系中激活cGMP产生的效力和效能分别比LGN高100倍和1000倍。在离体灌注的大鼠肾脏中，给药60分钟后，LGN(Ser)刺激钠排泄分数从24.8±3.0%最大增加到36.3±3.3%，并使尿流从0.15±0.01增加到0.24±0.01 ml·g⁻¹·min⁻¹。LGN(Ser)（0.69 microM）还使钾排泄分数从27.3±2.3%增加到38.0±3.0%，氯排泄分数从26.1±0.8%增加到43.5±1.9%。LGN(Ser)使尿中cGMP排泄量从1.00±0.04增加到9.28±1.14 pmol/ml，增加了9倍，而给药后cAMP水平没有变化。这些发现表明，与天然LGN一样含有单一二硫键的LGN(Ser)可激活T84和OK细胞中的鸟苷酸环化酶 - C（GC - C）受体，并且可能对研究体内激活GC - C的生理重要性非常有帮助。LGN(Ser)在离体灌注的肾脏中也表现出与先前观察到的负鼠尿鸟苷林相当的完全活性，这表明LGN在肾功能中具有生理作用。因此，单个氨基酸取代增强了LGN的活性和效力。
