Clawitter J, Trout W E, Burke M G, Araghi S, Roberts R M
Department of Biochemistry, University of Missouri, Columbia 65211.
J Biol Chem. 1990 Feb 25;265(6):3248-55.
Two-dimensional polyacrylamide gel electrophoresis has revealed the presence of a group of relatively acidic proteins of molecular weight about 22,000 in the uterine flushings of pseudopregnant pigs. The proteins have been purified by a combination of gel filtration chromatography and high performance anion-exchange chromatography and shown to bind both [3H] retinol and [3H]retinoic acid. At least four protein peaks that bound retinoids could be detected in the uterine secretions of a single pig. The ion-exchange procedure also allowed the retinol-free apoproteins to be separated from the holoforms that had associated ligand. Amino acid sequencing of the NH2 termini of polypeptides within three of the peaks revealed the presence of proteins with some degree of sequence identity to serum retinol-binding proteins (RBP). The most basic polypeptides showed the least similarity (about 30% identity), while the most acidic isoform analyzed shared about 70% sequence identity with the NH2 terminus of human serum RBP. Western blotting procedures employing an antiserum raised against the most basic isoforms showed that the amount of retinol-binding protein in uterine secretions increased markedly in ovariectomized animals in response to long term progesterone treatment. These proteins appear to form part of the uterine histotroph thought to be essential for nourishment of the conceptuses during pregnancy. A simple three-step procedure for purifying retinol-binding protein from pig serum is also described. The NH2-terminal sequence of this RBP is similar to that of human RBP but different from those of the uterine forms. The study suggests that a family of RBP, distinct from the serum form, is secreted by the uterine endometrium of the pig in response to progesterone.
二维聚丙烯酰胺凝胶电泳显示，在假孕母猪的子宫冲洗液中存在一组分子量约为22,000的相对酸性蛋白质。这些蛋白质通过凝胶过滤色谱法和高效阴离子交换色谱法相结合的方法进行了纯化，并显示出能结合[3H]视黄醇和[3H]视黄酸。在一头母猪的子宫分泌物中可检测到至少四个结合类视黄醇的蛋白峰。离子交换程序还能将无视黄醇的脱辅基蛋白与结合了配体的全蛋白形式分离。对三个峰内多肽的NH2末端进行氨基酸测序，结果显示存在与血清视黄醇结合蛋白（RBP）具有一定程度序列同一性的蛋白质。最碱性的多肽显示出的相似性最低（约30%同一性），而分析的最酸性同工型与人类血清RBP的NH2末端共享约70%的序列同一性。采用针对最碱性同工型产生的抗血清进行的蛋白质印迹程序表明，在长期孕酮处理后，去卵巢动物子宫分泌物中视黄醇结合蛋白的量显著增加。这些蛋白质似乎构成了子宫组织营养物的一部分，被认为对孕期胚胎的营养至关重要。本文还描述了一种从猪血清中纯化视黄醇结合蛋白的简单三步程序。这种RBP的NH2末端序列与人类RBP相似，但与子宫形式的不同。该研究表明，猪子宫内膜会响应孕酮分泌一种不同于血清形式的RBP家族。
