Purification of IGF-I in Bovine Colostrum and its Application for Animal Production Research.

牛初乳中IGF-I的纯化及其在动物生产研究中的应用。

• 批准号: 63860036
• 负责人: SASAKI Yasuyuki
• 金额: $ 5.12万
• 依托单位: Tohoku University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B).
• 财政年份: 1988
• 资助国家: 日本
• 起止时间: 1988 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-63860036/
• 关键词: Bovine; Colostrum; IGF-I; Purification

Insulin-like growth factor-I (IGF-I) is a biologically active peptide consisting of 70 amino acids (mol wt. 7,649). Its chemical structure is homologous to proinsulin. The primary structure of IGF-I are almost identical in mammalian species tested (human, rat, mouse, porcine, bovine, ovine).Biological action of IGF-I, firstly found as sulfation factor, has been shown by recent investigations to stimulate proliferation and differentiation of various cell types. Growth hormone stimulates the secretion of IGF-I, then IGF-I stimulates the growth of young animals through accelerating bone elongation and protein accumulation. On this point IGF-I has been recognized to be of importance for animal production. We showed that the concentration (0.5mg/l) of IGF-I in bovine colostrum exceeded that in blood plasma which has been used as a biological material to extract IGF-I, suggesting availability of colostrum to be an useful material for purifying IGF-I. Colostrum was collected on the day of par … More turition. Chymosin was added to colostrum. After incubation at 37 C, the lipid- and casein-free supernatant was obtained by centrifugation. Acid-ethanol mixture (12.5% HC1, 87.5% EtOH) was added to dissociate IGF-I from its binding protein, followed by further addition of cold acetone to get acetone powder. We used the acetone powder as starting material. The powder was dissolved in 1M acetic acid solution to apply to sephadex G-50 column. Eluate of IGF-I immunoreactive fractions, containing 0.7 ug IGF-I/mg protein, was lyophilized and gel filtration procedure by TSKG3000SW was performed. The TSK column was eluted with 1M acetic acid, IGF-I being purified to 1.6 ug/mg protein. The lyophilized sample was dissolved with 20% acetonitrile in 0.05% TFA, and applied to the first ODP-50 reverse-phase HPLC. IGF-I immunoreactivity was eluted by a two-step linear gradient to 80% acetonitrile. IGF-I concentration of the eluate was 32 ug/mg protein. IGF-I was further purified by the second ODP-50 reverse phase HPLC. The bioassay using rat myoblast subculture cells clearly showed IGF-I bioactivity in the samples eluted from Sephadex and TSK gel column. Less

胰岛素样生长因子-I（IGF-I）是由70个氨基酸（mol wt. 7，649）。其化学结构与胰岛素原同源。IGF-I的一级结构在所测试的哺乳动物物种（人、大鼠、小鼠、猪、牛、羊）中几乎是相同的。IGF-I的生物学作用首先被发现为硫酸化因子，最近的研究已显示其刺激各种细胞类型的增殖和分化。生长激素刺激IGF-I的分泌，IGF-I通过促进骨骼的伸长和蛋白质的积累来促进幼龄动物的生长。在这一点上，IGF-I已被公认为对动物生产的重要性。结果表明，牛初乳中IGF-I的浓度（0.5mg/l）超过了血浆中IGF-I的浓度，表明牛初乳是一种有效的纯化IGF-I的材料。在par当天收集初乳 ...更多信息 turition。将凝乳酶添加到初乳中。在37 ℃下孵育后，通过离心获得不含脂质和酪蛋白的上清液。加入酸-乙醇混合物（12.5%HC1，87.5%EtOH）以使IGF-I从其结合蛋白解离，随后进一步加入冷丙酮以得到丙酮粉末。我们使用丙酮粉末作为起始材料。将粉末溶解于1 M乙酸溶液中，应用于SephadexG-50柱。将含有0.7 μ g IGF-I/mg蛋白质的IGF-I免疫反应性级分的洗脱液冻干，并通过TSKG 3000 SW进行凝胶过滤程序。用1 M乙酸洗脱TSK柱，将IGF-1纯化至1.6 μ g/mg蛋白质。将冻干样品用20%乙腈的0.05% TFA溶液溶解，并应用于第一个ODP-50反相HPLC。IGF-I免疫反应性通过两步线性梯度至80%乙腈洗脱。IGF-I浓度为32微克/毫克蛋白质。通过第二次ODP-50反相HPLC进一步纯化IGF-I。用大鼠成肌细胞传代培养细胞进行的生物活性测定清楚地显示了从Sephadex和TSK凝胶柱洗脱的样品中IGF-I的生物活性。少

项目成果

小田伸一: 第80回日本畜産学会大会講演要旨集. 185 (1988)

Shinichi Oda：日本动物科学学会第 80 届年会摘要 185（1988）。

S.Oda: "Periparturient changes in concentration of insulin like growth factorーI in bovine colostrum and in plasma of dairy cows and neonatal calves" AsianーAustralasian J.Anim.Sci.2. 210-211 (1989)

S.Oda：“牛初乳以及奶牛和新生犊牛血浆中胰岛素样生长因子-I 浓度的围产期变化”亚洲-澳大利亚 J.Anim.Sci.2 (1989)。

小田伸一: "ウシの分娩に伴う血漿および初乳IGF-I、GH、インスリン、グルカゴン濃度変動について" 第80回日本畜産学会大会講演要旨集. 185 (1988)

Shinichi Oda：“奶牛产犊期间血浆和初乳 IGF-I、GH、胰岛素和胰高血糖素浓度的变化”日本动物科学学会第 80 届年会摘要 185 (1988)。

Oda S.: "Insulin like growth factor-I, GH, insulin and glucagon concentrations in bovine colostrum and in plasma of dairy cows and neonatal calves around parturition. (in Japanese)" Bulletin of The Exp. Farm, Tohoku Univ.5. 63-66 (1989)

Oda S.：“牛初乳以及分娩前后奶牛和新生犊牛血浆中的胰岛素样生长因子-I、GH、胰岛素和胰高血糖素浓度。（日语）”Exp 公告。

Oda S.: "Insulin like growth factor-I,GH,insulin and glucagon concentrations in bovine colostrum and in plasma of dairy cows and neonatal calves around parturition" Comp.Biochem.Physiol.94A. 805-808 (1989)

Oda S.：“牛初乳以及分娩前后奶牛和新生犊牛血浆中的胰岛素样生长因子-I、GH、胰岛素和胰高血糖素浓度”Comp.Biochem.Physiol.94A。