CLONING AND EXPRESSION OF CDNA FOR A NA/PI COTRANSPORT SYSTEM OF KIDNEY CORTEX
CLONING AND EXPRESSION OF CDNA FOR A NA/PI COTRANSPORT SYSTEM OF KIDNEY CORTEX
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DOI：
10.1073/pnas.88.21.9608
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发表时间：
1991-11-01
影响因子：
11.1
通讯作者：
MURER, H
中科院分区：
文献类型：
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作者：
WERNER, A;MOORE, ML;MURER, H
以非洲爪蟾卵母细胞为表达系统，从兔肾皮质cDNA文库中筛选了磷酸盐钠依赖转运蛋白（P（i））的表达。最终分离出一个单一的克隆（命名为NaP（i）-1），其刺激Na/P（i）共转运的表达几乎等于总mRNA的700倍。Na/P（i）协同转运蛋白的预测序列由465个氨基酸组成（相对分子质量，51，797）;亲水性特征预测表明有6个（可能是8个）跨膜片段。在胰腺微粒体的存在下，NaPi-1/互补RNA的体外翻译表明NaPi-1是一种糖基化蛋白;氨基酸序列中存在四个潜在的N-糖基化位点。北方印迹分析表明，NaPi-1/mRNA存在于肾皮质和肝脏;没有杂交信号与mRNA从其他组织（包括小肠）。NaPi-1/互补RNA表达的Na/P（i）共转运的动力学分析表明，其特征（钠相互作用）与在皮质顶膜中观察到的特征相似。5个氨基酸残基（Gly 342/Ala 381 XaaXaaXaaXaaLeu 386 XaaXaaXaaPro 390 Arg 391）的比对与针对Na依赖性转运系统提出的基序一致。我们的结论是，我们已经克隆了一个cDNA的Na/P（i）共转运系统存在于兔肾皮质。
A cDNA library from rabbit kidney cortex was screened for expression of Na-dependent transport of phosphate (P(i)) using Xenopus laevis oocytes as an expression system. A single clone was eventually isolated (designated NaP(i)-1) that stimulated expression of Na/P(i) cotransport almost-equal-to 700-fold compared to total mRNA. The predicted sequence of the Na/P(i) cotransporter consists of 465 amino acids (relative molecular mass, 51,797); hydropathy profile predictions suggest six (possibly eight) membrane-spanning segments. In vitro translation of NaPi-1/complementary RNA in the presence of pancreatic microsomes indicated NaPi-1 to be a glycosylated protein; four potential N-glycosylation sites are present in the amino acid sequence. Northern blot analysis demonstrated the presence of NaPi-1/mRNA in kidney cortex and liver; no hybridization signal was obtained with mRNA from other tissues (including small intestine). Kinetic analysis of Na/P(i) cotransport expressed by NaPi-1/complementary RNA demonstrated characteristics (sodium interaction) similar to those observed in cortical apical membranes. The alignment of 5 amino acid residues (Gly342/Ala381XaaXaaXaaXaaLeu386XaaXaaXaaPro390Arg391) is consistent with a motif proposed for Na-dependent transport systems. We conclude that we have cloned a cDNA for a Na/P(i) cotransport system present in rabbit kidney cortex.