A RABBIT AUTOANTIBODY SPECIFIC FOR THE 46-KDA FORM OF 2',3'-CYCLIC NUCLEOTIDE 3'-PHOSPHODIESTERASE
A RABBIT AUTOANTIBODY SPECIFIC FOR THE 46-KDA FORM OF 2',3'-CYCLIC NUCLEOTIDE 3'-PHOSPHODIESTERASE
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DOI：
10.1111/j.1471-4159.1992.tb10059.x
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发表时间：
1992-05-01
影响因子：
4.7
通讯作者：
QUARLES, RH
中科院分区：
文献类型：
--
作者：
MOLLER, JR;RAMASWAMY, SG;QUARLES, RH
在明显正常的兔子血清中出现的一种自身抗体对大鼠大脑中的髓鞘和少突胶质细胞进行免疫细胞化学染色，研究表明该抗体与许多物种中的2 '，3'-环核苷酸3 '-磷酸二酯酶（CNP）的46-kDa亚型发生特异性反应（EC 3.1.4.37）。通过比较自身抗体与充分表征的抗CNP抗血清的蛋白质印迹免疫染色，确定酶的较短亚型（CNP 1）为抗原。与自身抗体反应的46-kDa抗原在二维凝胶上显示出与CNP的小亚型相同的M（r）和pI，并显示出在纯化的CNS髓鞘中相似的富集。自身抗体对CNP 1具有非常高的亲和力，并且能够检测外周神经、脾脏、肾上腺、胰腺、睾丸和肠中非常少量的这种酶。通过酶联免疫吸附测定法测试自身抗体与合成肽的反应性，结果显示其与对应于CNP 1的N-末端的N-乙酰化十肽反应，但如果肽未被乙酰化或如果乙酰基被棕榈基取代则不反应。由于选择性剪接，CNP 2与CNP 1的不同之处在于蛋白质N末端的20个氨基酸延伸，与CNP 2缺乏反应性可能是由于不存在作为表位一部分的N-乙酰基部分和/或通过多肽链的延伸阻断抗体与十肽的结合。这种针对CNP较短亚型的高度特异性抗体可用于确定该酶两种亚型的位置或功能可能差异的研究。
An autoantibody occurring in the serum of an apparently normal rabbit that immunocytochemically stains myelin sheaths and oligodendrocytes in rat brain was shown to react specifically with the 46-kDa isoform of 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) (EC 3.1.4.37) in a number of species. Identification of the shorter isoform of the enzyme (CNP1) as the antigen was achieved by comparing the immunostaining of Western blots by the autoantibody with that of a well-characterized anti-CNP antiserum. The 46-kDa antigen reacting with the autoantibody exhibited the same M(r) and pI as the small isoform of CNP on two-dimensional gels and showed a similar enrichment in purified CNS myelin. The autoantibody has very high affinity for CNP1 and is capable of detecting the very low amounts of this enzyme in peripheral nerve, spleen, adrenal gland, pancreas, testis, and intestine. Testing the reactivity of the autoantibody with synthetic peptides by enzyme-linked immunosorbent assay revealed that it reacted with the N-acetylated decapeptide corresponding to the N-terminus of CNP1, but did not react if the peptide was not acetylated or if the acetyl group was replaced with a palmityl group. The lack of reactivity with CNP2, which differs from CNP1 by a 20-amino acid extension at the N-terminus of the protein as a result of alternative splicing, may be due to the absence of the N-acetyl moiety that is part of the epitope and/or blocking of antibody binding to the decapeptide by extension of the polypeptide chain. This highly specific antibody for the shorter isoform of CNP may be useful in studies to establish possible differences in the location or function of the two isoforms of this enzyme.