title: Calpain-resistant fragment(s) of alpha-synuclein regulates the synuclein-cleaving activity of 20S proteasome.
abstract: Alpha-synuclein is a pathological component of Parkinson's disease by participating in Lewy body formation. Imbalance in protein turnover could result in the abnormal protein aggregation responsible for eventual neuronal cell death. This in vitro digestion study showed that both m-calpain and 20S proteasome preferentially hydrolyzed the N-terminal half of alpha-synuclein, which made the hydrophobic NAC and following acidic C-terminal region resistant against the proteolyses. Since the acidic C-terminal region contains the PEST segment-a protein degradation signal enriched with amino acids of proline (P), glutamate (E), serine (S), and threonine (T)-, the PEST segment has not been processed or even required for the proteolyses. Alpha-synuclein would be recognized primarily by m-calpain since the common substrate was processed by m-calpain five times more effectively than 20S proteasome with k(cat)/K(m) of 1.64 x 10(4)M(-1)s(-1) and 0.32 x 10(4) M(-1)s(-1), respectively. The N-terminally truncated protease-resistant C-terminal fragment of alpha-syn61-140 was demonstrated to stimulate the 20S proteasome-mediated breakdown of alpha-synuclein and its mutant forms of Ala53Thr and Ala30Pro. The stimulation for Ala53Thr, however, was noticeably less efficient than those for the other proteins, which might support the previous observation of the prolonged intracellular life span of Ala53Thr by 1.5-fold compared to that of wild-type form. We have hypothesized that the N-terminally truncated C-terminal fragment derived from the abundant alpha-synuclein through intracellular proteolyses could be involved in various physiological or pathological effects which might be related to the formation of abnormal protein aggregation and subsequent neuronal degeneration by influencing the intracellular protein turnover or directly participating in the aggregate formation.
authors.affiliation: School of Chemical and Biological Engineering, College of Engineering, Seoul National University, Seoul 151-744, Republic of Korea.
authors.forename: Hyun Jin
authors.initials: HJ
authors.lastname: Kim
authors.forename: Daekyun
authors.initials: D
authors.lastname: Lee
authors.forename: Choong-Hwan
authors.initials: CH
authors.lastname: Lee
authors.forename: Kwang Chul
authors.initials: KC
authors.lastname: Chung
authors.forename: Jongsun
authors.initials: J
authors.lastname: Kim
authors.forename: Seung R
authors.initials: SR
authors.lastname: Paik
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000596:Amino Acids / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002154:Calpain / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005971:Glutamates / Q000235:genetics / Q000378:metabolism; D006868:Hydrolysis; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011392:Proline / Q000235:genetics / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D012694:Serine / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D013552:Swine; D013912:Threonine / Q000235:genetics / Q000378:metabolism; D051844:alpha-Synuclein / Q000235:genetics / Q000378:metabolism*
chemical_list: D000596:Amino Acids; D005971:Glutamates; D051844:alpha-Synuclein; D013912:Threonine; D012694:Serine; D011392:Proline; D002154:Calpain; D046988:Proteasome Endopeptidase Complex