Gunnersen D, Haley B
Department of Biochemistry, College of Pharmacy, University of Kentucky, Lexington 40536-0084.
Proc Natl Acad Sci U S A. 1992 Dec 15;89(24):11949-53. doi: 10.1073/pnas.89.24.11949.
In this report, 8- and 2-azidoadenosine 5'-[gamma-32P]triphosphate were used to examine cerebrospinal fluid (CSF) samples for the presence of an ATP binding protein unique to individuals with Alzheimer disease (AD). A 42-kDa ATP binding protein was found in the CSF of AD patients that is not observed in CSF from normal patients or other neurological controls. The photolabeling is saturated with 30 microM 2-azidoadenosine 5'-[gamma-32P]triphosphate. Photoinsertion can be totally prevented by the addition of 25 microM ATP. Photoinsertion of 2-azidoadenosine 5'-triphosphate into the protein is only weakly protected by other nucleotides such as ADP and GTP, indicating that this is a specific ATP binding protein. A total of 83 CSF samples were examined in a blind manner. The 42-kDa protein was detected in 38 of 39 AD CSF samples and in only 1 of 44 control samples. This protein was identified as glutamine synthetase [GS; glutamate-ammonia ligase; L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2] based on similar nucleotide binding properties, comigration on two-dimensional gels, reaction with a polyclonal anti-GS antibody, and the presence of significant GS enzyme activity in AD CSF. In brain, GS plays a key role in elimination of free ammonia and also converts the neurotransmitter and excitotoxic amino acid glutamate to glutamine, which is not neurotoxic. The involvement of GS, if any, in the onset of AD is unknown. However, the presence of GS in the CSF of terminal AD patients suggests that this enzyme may be a useful diagnostic marker and that further study is warranted to determine any possible role for glutamate metabolism in the pathology of AD.
在本报告中，使用8 - 叠氮腺苷5'-[γ-32P]三磷酸和2 - 叠氮腺苷5'-[γ-32P]三磷酸来检测脑脊液（CSF）样本，以确定阿尔茨海默病（AD）患者所特有的一种ATP结合蛋白的存在情况。在AD患者的脑脊液中发现了一种42 kDa的ATP结合蛋白，而在正常患者或其他神经学对照的脑脊液中未观察到。用30 microM 2 - 叠氮腺苷5'-[γ-32P]三磷酸时光标记达到饱和。加入25 microM ATP可完全阻止光插入。2 - 叠氮腺苷5'-三磷酸插入该蛋白的过程仅受到其他核苷酸如ADP和GTP的微弱保护，表明这是一种特异性的ATP结合蛋白。以盲法检测了总共83份脑脊液样本。在39份AD脑脊液样本中的38份以及44份对照样本中的1份中检测到了这种42 kDa的蛋白。基于相似的核苷酸结合特性、在二维凝胶上的共迁移、与多克隆抗GS抗体的反应以及AD脑脊液中存在显著的GS酶活性，该蛋白被鉴定为谷氨酰胺合成酶[GS；谷氨酸 - 氨连接酶；L - 谷氨酸：氨连接酶（形成ADP），EC 6.3.1.2]。在大脑中，GS在清除游离氨方面起关键作用，还将神经递质和具有兴奋性毒性的氨基酸谷氨酸转化为无神经毒性的谷氨酰胺。GS在AD发病过程中（若有）的作用尚不清楚。然而，晚期AD患者脑脊液中存在GS表明该酶可能是一种有用的诊断标志物，有必要进一步研究以确定谷氨酸代谢在AD病理过程中的任何可能作用。
