Expression of separate isoforms of human tau protein: correlation with the tau pattern in brain and effects on tubulin polymerization.
We have expressed six previously cloned isoforms of human microtubule-associated tau protein in Escherichia coli and purified them to homogeneity in a biologically active form. They range from 352 to 441 amino acids in length and differ from each other by the presence of three or four tandem repeats in the carboxy-terminal half and by the presence or absence of 29 or 58 amino acid inserts in the amino-terminus. When mixed together they gave a set of six bands on SDS-PAGE gels with apparent molecular weights of 48-67 kd and with a characteristic pattern of spacings. Four of these bands aligned with the major tau bands found in adult human cerebral cortex following perchloric acid extraction and alkaline phosphatase treatment. They consisted of isoforms with three repeats and no insertions, four repeats and no amino-terminal insertions and three- and four-repeat containing isoforms with the 29 amino acid insertion. In fetal human brain extracts treated with alkaline phosphatase one of the two major tau bands aligned with the three-repeat containing isoform with no insertions, whereas the molecular nature of the second major tau band remains to be established. The recombinant tau isoforms were biologically active at micromolar concentrations, as assessed by their ability to promote microtubule assembly. The rates of assembly were 2.5-3.0 times faster for isoforms containing four repeats when compared with three-repeat containing isoforms, with no significant contribution by the amino-terminal insertions.
Goedert
M
M
Medical Research Council Laboratory of Molecular Biology, Cambridge, UK.
Jakes
R
R
D000368:Aged; D000469:Alkaline Phosphatase / Q000494:pharmacology; D000544:Alzheimer Disease / Q000473:pathology; D000595:Amino Acid Sequence; D001921:Brain / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D004926:Escherichia coli / Q000032:analysis / Q000254:growth & development; D006801:Humans; D007536:Isomerism; D008869:Microtubule-Associated Proteins / Q000096:biosynthesis / Q000737:chemistry*; D008875:Middle Aged; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D012326:RNA Splicing; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D014404:Tubulin / Q000096:biosynthesis / Q000235:genetics*; D016875:tau Proteins
D008869:Microtubule-Associated Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014404:Tubulin; D016875:tau Proteins; D000469:Alkaline Phosphatase