Hogue-Angeletti R A, Frazier W A, Jacobs J W, Niall H D, Bradshaw R A
Biochemistry. 1976 Jan 13;15(1):26-34. doi: 10.1021/bi00646a005.
The nerve growth factor (NGF) from Naja naja (cobra) venom has been purified and its structure compared to the NGF from mouse submaxillary gland. A two-step purification procedure has been devised, consisting of a gel filtration step in 1 M acetic acid followed by chromatography of the active pool on carboxymethylcellulose at pH 5. The molecular weight of the native protein was found to be 28000, and this value was reduced by approximately one-half under denaturing conditions. These values are comparable to those obtained for mouse 2.5S or betaNGF. Tryptic peptide maps of S-[14C]carboxymethyl NGF gave the number of labeled peptides expected for a structure composed of two identical or very similar subunits. Thus, the quaternary structures of mouse and cobra NGF are the same. Cyanogen bromide (CNBr) treatment of Naja naja NGF produced three fragments, of which two were purified to homogeneity. These fragments and the whole protein were analyzed in the automated protein Sequencer. The amino-terminal CNBr fragment of the protein was also subjected to digestion by thermolysin and the resultant peptides were purified and characterized. These data plus those from the characterization of the tryptic peptides provided the basis of the construction of a tentative primary structure of Naja naja NGF which is approximately 60% identical with mouse NGF.
已从眼镜蛇毒液中纯化出神经生长因子（NGF），并将其结构与小鼠颌下腺的NGF进行了比较。设计了一种两步纯化程序，包括在1 M乙酸中进行凝胶过滤步骤，然后在pH 5的条件下将活性组分在羧甲基纤维素上进行色谱分离。发现天然蛋白质的分子量为28000，在变性条件下该值降低了约一半。这些值与从小鼠2.5S或βNGF获得的值相当。S-[14C]羧甲基NGF的胰蛋白酶肽图谱给出了由两个相同或非常相似的亚基组成的结构所预期的标记肽数量。因此，小鼠和眼镜蛇NGF的四级结构相同。用溴化氰（CNBr）处理眼镜蛇NGF产生了三个片段，其中两个被纯化至同质。这些片段和完整蛋白质在自动蛋白质测序仪中进行了分析。该蛋白质的氨基末端CNBr片段也用嗜热菌蛋白酶进行了消化，所得肽段被纯化并进行了表征。这些数据以及来自胰蛋白酶肽表征的数据为构建眼镜蛇NGF的初步一级结构提供了基础，该结构与小鼠NGF约60%相同。
