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    • 6. 发明申请
    • MANUFACTURE AND USE OF POLYPEPTIDES TAGGED USING BINDING MOLECULES
    • 使用结合分子标记的多肽的制备和使用
    • WO1995012672A1
    • 1995-05-11
    • PCT/GB1994002420
    • 1994-11-04
    • MEDICAL RESEARCH COUNCILNERI, DarioWINTER, Gregory, PaulDE LALLA, Claudia
    • MEDICAL RESEARCH COUNCIL
    • C12N15/12
    • C07K14/4728C07K14/43568C07K14/47C07K2319/00G01N33/68
    • Molecules comprise (i) a polypeptide (such as calmodulin) which has calcium-dependent binding affinity for ligand and (ii) another polypeptide, the polypeptides preferably being joined by a peptide bond and produced by recombinant expression from a gene fusion. The molecules are useful in detection, immobilisation, targeting and purification, cell-labelling, and band-shift assays for determining binding of a member of a specific binding pair (sbp) for complementary sbp member. For purposes of band-shift assays, polypeptide (i) need not have calcium-dependent binding affinity for a ligand, but should have a dissociation constant for a ligand of 10nM or less, measured at a pH of between 6 and 9 at 20 DEG C. In an alternative embodiment, a calmodulin-binding polypeptide which is, or is derived from, mastoparan is joined, as polypeptide (i) instead of a binding polypeptide, to the other polypeptide.
    • 分子包含(i)对配体具有钙依赖性结合亲和力的多肽(例如钙调素)和(ii)另一种多肽,所述多肽优选通过肽键连接并通过基因融合体的重组表达产生。 这些分子可用于检测,固定,靶向和纯化,细胞标记和带状移位测定,以确定用于互补sbp成员的特异性结合对(sbp)的成员的结合。 为了频移测定的目的,多肽(i)不需要对配体具有钙依赖性结合亲和力,但是对于配体的解离常数为10nM或更低,在pH为6至9的20℃下测量。 C.在替代实施方案中,将多肽(i)代替结合多肽连接到另一多肽中,所述钙调蛋白结合多肽或其衍生自mastoparan的钙调蛋白结合多肽与其他多肽相连接。