Synapsin I is a major endogenous substrate for protein L-isoaspartyl methyltransferase in mammalian brain.
The accumulation of potentially deleterious L-isoaspartyl linkages in proteins is prevented by the action of protein L-isoaspartyl O-methyltransferase, a widely distributed enzyme that is particularly active in mammalian brain. Methyltransferase-deficient (knock-out) mice exhibit greatly increased levels of isoaspartate and typically succumb to fatal epileptic seizures at 4-10 weeks of age. The link between isoaspartate accumulation and the neurological abnormalities of these mice is poorly understood. Here, we demonstrate that synapsin I from knock-out mice contains 0.9 +/- 0.3 mol of isoaspartate/mol of synapsin, whereas the levels in wild-type and heterozygous mice are undetectable. Transgenic mice that selectively express methyltransferase only in neurons show reduced levels of synapsin damage, and the degree of reduction correlates with the phenotype of these mice. Isoaspartate levels in synapsin from the knock-out mice are five to seven times greater than those in the average protein from brain cytosol or from a synaptic vesicle-enriched fraction. The isoaspartyl sites in synapsin from knock-out mice are efficiently repaired in vitro by incubation with purified methyltransferase and S-adenosyl-L-methionine. These findings demonstrate that synapsin I is a major substrate for the isoaspartyl methyltransferase in neurons and suggest that isoaspartate-related alterations in the function of presynaptic proteins may contribute to the neurological abnormalities of mice deficient in this enzyme.
Reissner
Kathryn J
KJ
Department of Molecular Biology and Biochemistry, University of California, Irvine, California 92697-3900, USA.
Paranandi
Mallik V
MV
Luc
Trang M
TM
Doyle
Hester A
HA
Mamula
Mark J
MJ
Lowenson
Jonathan D
JD
Aswad
Dana W
DW
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D001921:Brain / Q000201:enzymology* / Q000378:metabolism*; D002147:Calmodulin / Q000032:analysis / Q000302:isolation & purification; D002417:Cattle; D002458:Cell Fractionation; D016000:Cluster Analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006579:Heterozygote; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008745:Methylation; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase / Q000032:analysis / Q000737:chemistry / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions; D013379:Substrate Specificity; D016704:Synapsins / Q000302:isolation & purification / Q000378:metabolism*; D014357:Trypsin / Q000494:pharmacology
D002147:Calmodulin; D007527:Isoenzymes; D016704:Synapsins; D000255:Adenosine Triphosphate; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase; D014357:Trypsin