Varro A, Nemeth J, Bridson J, Lee C, Moore S, Dockray G J
Department of Physiology, University of Liverpool, United Kingdom.
J Biol Chem. 1990 Dec 15;265(35):21476-81.
Post-translational processing of the precursor for rat gastrin yields products that include peptides phosphorylated at Ser96, amidated at Phe92, and sulfated at Tyr87 or Tyr103. The phosphorylation site is immediately adjacent to the processing point that gives rise to the biologically active amidated gastrins. We have examined changes in post-translational processing which occur in gastrin cells from rats that are physiologically stimulated (by feeding) or unstimulated (by fasting). Peptides were identified using site-directed radioimmunoassays and chromatographic systems that resolve phosphorylated, amidated, and sulfated progastrin products, including intermediates generated prior to amidation (i.e. C-terminal glycine-extended variants). Assays for Phe92-amidated peptides and for the C-terminal tryptic fragment of progastrin indicated decreases in the total tissue concentrations of immunoreactive peptide with fasting; in contrast, the tissue concentrations of glycine-extended biosynthetic intermediates were similar in fasted and fed rats. Taken together the data suggest a relative failure in amidation mechanisms in unstimulated cells. The endopeptidase cleavage of progastrin was not influenced significantly by fasting. However, the phosphorylation of peptide products containing Ser96 was depressed significantly in fasted rats. The proportions of amidated peptides sulfated at Tyr87 were generally lower than their corresponding glycine-extended biosynthetic precursors, but in both cases the proportion of peptide in the sulfated form was lower than for peptides sulfated at Tyr103. Feeding did not change the sulfation of amidated heptadecapeptide gastrin or its glycine-extended variant. The results suggest that the mechanisms determining phosphorylation and amidation of progastrin-related peptides depend on the patterns of stimulation of gastrin cells. The observation that decreased phosphorylation is associated with a failure to produce active amidated products is consistent with a regulatory function for phosphorylation in gastrin production.
大鼠胃泌素前体的翻译后加工产生的产物包括在Ser96处磷酸化、在Phe92处酰胺化以及在Tyr87或Tyr103处硫酸化的肽段。磷酸化位点紧邻产生生物活性酰胺化胃泌素的加工位点。我们研究了生理刺激（喂食）或未刺激（禁食）的大鼠胃泌素细胞中发生的翻译后加工变化。使用定点放射免疫测定法和色谱系统鉴定肽段，该色谱系统可分离磷酸化、酰胺化和硫酸化的前胃泌素产物，包括酰胺化之前产生的中间体（即C末端甘氨酸延伸变体）。对Phe92酰胺化肽段和前胃泌素C末端胰蛋白酶片段的测定表明，禁食时免疫反应性肽段的总组织浓度降低；相反，禁食和喂食大鼠中甘氨酸延伸生物合成中间体的组织浓度相似。综合这些数据表明，未刺激细胞中的酰胺化机制相对失效。前胃泌素的内肽酶切割不受禁食的显著影响。然而，禁食大鼠中含有Ser96的肽产物的磷酸化明显降低。Tyr87硫酸化的酰胺化肽段比例通常低于其相应的甘氨酸延伸生物合成前体，但在这两种情况下，硫酸化形式的肽段比例均低于Tyr103硫酸化的肽段。喂食并未改变酰胺化十七肽胃泌素或其甘氨酸延伸变体的硫酸化。结果表明，决定前胃泌素相关肽段磷酸化和酰胺化的机制取决于胃泌素细胞的刺激模式。磷酸化降低与无法产生活性酰胺化产物相关的观察结果与磷酸化在胃泌素产生中的调节功能一致。
