Modulation of procaspase-7 self-activation by PEST amino acid residues of the N-terminal prodomain and intersubunit linker

Autor(es)

Alves, Juliano ^{[1, 2]} ; Garay-Malpartida, Miguel ^[3] ; Occhiucci, Joao M. ^[1] ; Belizario, Jose E. ^[1]

Afiliação do(s) autor(es)

^[1] Inst Biomed Sci, Dept Pharmacol, Ave Lineu Prestes 1524, BR-05508900 Sao Paulo, SP - Brazil

^[2] Promega Corp, 2800 Woods Hollow Rd, Madison, WI 53711 - USA

^[3] Univ Sao Paulo, Sch Arts Commun & Humanity, Rua Arlindo Bettio 1000, BR-03828000 Sao Paulo, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: BIOCHEMISTRY AND CELL BIOLOGY; v. 95, n. 6, p. 634-643, DEC 2017.
• Citações Web of Science: 0

Resumo

Procaspase-7 zymogen polypeptide is composed of a short prodomain, a large subunit (p20), and a small subunit (p10) connected to an intersubunit linker. Caspase-7 is activated by an initiator caspase-8 and -9, or by autocatalysis after specific cleavage at IQAD(198)down arrow S located at the intersubunit linker. Previously, we identified that PEST regions made of amino acid residues Pro (P), Glu (E), Asp (D), Ser (S), Thr (T), Asn (N), and Gln (Q) are conserved flanking amino acid residues in the cleavage sites within a prodomain and intersubunit linker of all caspase family members. Here we tested the impact of alanine substitution of PEST amino acid residues on procaspase-7 proteolytic self-activation directly in Escherichia coli. The p20 and p10 subunit cleavage were significantly delayed in double caspase-7 mutants in the prodomain (N18A/P26A) and intersubunit linker (S199A/P201A), compared with the wild-type caspase-7. The S199A/P201A mutants effectively inhibited the p10 small subunit cleavage. However, the mutations did not change the kinetic parameters (k(cat)/K-M) and optimal tetrapeptide specificity (DEVD) of the purified mutant enzymes. The results suggest a role of PEST-amino acid residues in the molecular mechanism for prodomain and intersubunit cleavage and caspase-7 self-activation. (AU)

• Processo FAPESP: 01/01000-7 - Análise da expressão de genes e proteínas e da atividade de enzimas caspases em células do músculo esquelético durante a caquexia induzida por tumores malignos e citocinas
• Beneficiário: José Ernesto Belizario
• Modalidade de apoio: Auxílio à Pesquisa - Regular