Epand R M, Epand R F, Stafford A R, Orlowski R C
Department of Biochemistry, McMaster University Health Sciences Centre, Hamilton, Ontario, Canada.
J Med Chem. 1988 Aug;31(8):1595-8. doi: 10.1021/jm00403a019.
Salmon calcitonin has an amino acid sequences that would allow it to form an amphipathic helix from approximately residue 9 to residue 22. We have synthesized a number of analogues of this peptide hormone with deletions in the carboxyl terminus of this putative amphipathic helix. These analogues include deletions of single amino acid residues at positions 19, 20, 21, or 22 as well as deletions of progressively larger segments starting with residue 19 and including deletions of residues 19 and 20; 19, 20, and 21; or 19, 20, 21, and 22. There is a small decrease in the helical content of these analogues compared with the native hormone, both in the presence and absence of amphiphiles. However, the extent of formation of secondary structure, as measured by circular dichroism, is similar for these deletion sequences as it is for the native hormone. In all cases, there is a large increase in the helical content of the peptide in the presence of dimyristoylphosphatidylglycerol, lysolecithin, or sodium dodecyl sulfate. All of the analogues have hypocalcemic activity in vivo in rats, comparable to the native hormone, except for des-Leu19-salmon calcitonin, which is about twice as active as the unmodified hormone. With use of an in vitro assay of adenylate cyclase activation in purified rat kidney membranes, des-Tyr22-salmon calcitonin, des-Leu19,Gln20,Thr21-salmon calcitonin, and des-Leu19Gln20,Thr21,Thr22-salmon calcitonin exhibited about one-tenth the stimulatory activity of the native hormone. Des-Tyr22-sCT and des-Leu19,Gln20,Thr21,Tyr22-sCT were also tested for their activity in inhibiting prolactin release from isolated rat pituitary cells. Both of these analogues exhibited inhibitory activity. Thus, the region of residues 19-22 does not greatly affect either the conformational or the biological properties of salmon calcitonin.
鲑鱼降钙素具有一种氨基酸序列，使其能够从大约第9位残基到第22位残基形成一个两亲性螺旋。我们已经合成了许多这种肽激素的类似物，在这个假定的两亲性螺旋的羧基末端有缺失。这些类似物包括在第19、20、21或22位单氨基酸残基的缺失，以及从第19位残基开始逐渐更大片段的缺失，包括第19和20位残基的缺失；第19、20和21位残基的缺失；或第19、20、21和22位残基的缺失。与天然激素相比，无论有无两亲分子，这些类似物的螺旋含量都有小幅下降。然而，通过圆二色性测量的二级结构形成程度，这些缺失序列与天然激素相似。在所有情况下，在二肉豆蔻酰磷脂酰甘油、溶血卵磷脂或十二烷基硫酸钠存在下，肽的螺旋含量都有大幅增加。所有类似物在大鼠体内都具有降钙活性，与天然激素相当，除了去亮氨酸19 - 鲑鱼降钙素，其活性约为未修饰激素的两倍。使用纯化的大鼠肾膜中腺苷酸环化酶激活的体外测定法，去酪氨酸22 - 鲑鱼降钙素、去亮氨酸19、谷氨酰胺20、苏氨酸21 - 鲑鱼降钙素和去亮氨酸19、谷氨酰胺20、苏氨酸21、苏氨酸22 - 鲑鱼降钙素表现出约为天然激素刺激活性十分之一的活性。还测试了去酪氨酸22 - sCT和去亮氨酸19、谷氨酰胺20、苏氨酸21、酪氨酸22 - sCT在抑制分离的大鼠垂体细胞释放催乳素方面的活性。这两种类似物都表现出抑制活性。因此，第19 - 22位残基区域对鲑鱼降钙素的构象或生物学性质影响不大。
