Abnormal properties of creatine kinase in Alzheimer's disease brain: correlation of reduced enzyme activity and active site photolabeling with aberrant cytosol-membrane partitioning.
The report shows that Alzheimer's disease (AD) brain creatine kinase (CK) is modified such that the nucleotide binding site of CK is blocked and that abnormal partitioning of CK between the soluble and pellet fractions occurs. First, CK activity was 86% decreased in AD brain homogenates in comparison to age-matched controls. Secondly, over a 23.5 fold greater 32P photoincorporation of [alpha 32P]8N3ATP was observed into CK of control vs. AD samples. Also, a 7.4-fold increase of enzyme induced 32P incorporation was observed in controls vs. AD samples by incubation with [gamma 32P]ATP. Thirdly, Western blot analysis showed that CK copy numbers in the AD homogenate were decreased by less than 14% in comparison to controls. However, analysis showed that control supernatant and pellet fractions contained 10.3 and 0.4 times the CK copy number found in the corresponding AD fractions. 32P incorporation by both photolabeling and enzyme catalyzed incorporation of radiolabel followed CK activity and not CK copy number. Further, [alpha 32P]ADP and [gamma 32P]ATP incorporated 32P into control brain and purified brain CK equally well, indicating that a mechanism different from gamma-phosphoryl transfer is involved in the enzymatic incorporation of radiolabel. Also, the level of abnormal partitioning of CK into AD brain pellet correlated with the decreased [32P]8N3GTP photolabeling and abnormal partitioning of beta-tubulin, a protein known to be aberrantly modified in the AD brain. This indicates that a common chemistry is affecting both CK and tubulin in AD.
David
S
S
Department of Chemistry, University of Kentucky, Lexington, KY 40506-0055, USA.
Shoemaker
M
M
Haley
B E
BE
D000544:Alzheimer Disease / Q000201:enzymology*; D001665:Binding Sites; D001921:Brain / Q000201:enzymology*; D016022:Case-Control Studies; D002451:Cell Compartmentation; D003402:Creatine Kinase / Q000737:chemistry*; D003600:Cytosol / Q000648:ultrastructure*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007425:Intracellular Membranes / Q000648:ultrastructure*; D007527:Isoenzymes; D010766:Phosphorylation; D019920:Photoaffinity Labels; D012995:Solubility; D014404:Tubulin
D007527:Isoenzymes; D019920:Photoaffinity Labels; D014404:Tubulin; D003402:Creatine Kinase