In vitro analysis of tau phosphorylation sites and its biological activity.
OBJECTIVE
To explore the association between the abnormal phosphorylation sites found in Alzheimer disease (AD) tau and the inhibition of its biological activity.


METHODS
Ultracentrifugation, chromatography, manual Edman degradation and autosequence techniques were used to prepare and phosphorylate human recombinant tau, isolate and purify 32P tau peptides and determine phosphorylation sites.


RESULTS
Phosphorylation of tau by casein kinase 1 (CK-1), cyclic AMP-dependent protein kinase (PKA) and glycogen synthetase kinase 3 (GSK-3) separately inhibited its biological activity and the inhibition of this activity by (CSK-3 was significantly increased if tau was prephosphorylated by CK-1 or PKA. The most potent inhibition was seen by a combined phosphorylation of tau with PKA and GSK-3. The treatment of tau by PKA and GSK-3 combination induced phosphorylation of tau at Ser-195, Ser-198, Ser-199, Ser-202, Thr-205, Thr-231, Ser-235, Ser-262, Ser-356, Ser-404, whereas Thr-181, Ser-184, Ser-262, Ser-356 and Ser-400 were phosphorylated by GSK-3 alone under the same condition.


CONCLUSIONS
Phosphorylation of tau by PKA plus GSK-3 at Thr-205 might play a key role in tau pathology in AD.
Wang
Jianzhi
J
Tongji Medical College, Hua Zhong University of Science and Technology, Wuhan 430030.
Wang
Xiaochuan
X
Liu
Rong
R
Wang
Qun
Q
Grundke-Iqbal
Inge
I
Iqbal
Khalid
K
D000544:Alzheimer Disease / Q000378:metabolism; D001665:Binding Sites; D047388:Casein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008870:Microtubules / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D016875:tau Proteins / Q000378:metabolism*
D016875:tau Proteins; D011494:Protein Kinases; D047388:Casein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3