Montero-Morán Gabriela M, Sampedro José G, Saab-Rincón Gloria, Cervantes-González Miguel A, Huerta-Ocampo José Á, De León-Rodríguez Antonio, Barba de la Rosa Ana P
División Biología Molecular, IPICyT, Instituto Potosino de Investigación Científica y Tecnológica, Camino a la Presa San José 2055, Col. Lomas 4a. Sección, 78216, San Luis Potosí, SLP, Mexico.
Appl Biochem Biotechnol. 2015 Aug;176(8):2328-45. doi: 10.1007/s12010-015-1721-0. Epub 2015 Jul 1.
A novel Cu/ZnSOD from Amaranthus hypochondriacus was cloned, expressed, and characterized. Nucleotide sequence analysis showed an open reading frame (ORF) of 456 bp, which was predicted to encode a 15.6-kDa molecular weight protein with a pI of 5.4. Structural analysis showed highly conserved amino acid residues involved in Cu/Zn binding. Recombinant amaranth superoxide dismutase (rAhSOD) displayed more than 50 % of catalytic activity after incubation at 100 °C for 30 min. In silico analysis of Amaranthus hypochondriacus SOD (AhSOD) amino acid sequence for globularity and disorder suggested that this protein is mainly disordered; this was confirmed by circular dichroism, which showed the lack of secondary structure. Intrinsic fluorescence studies showed that rAhSOD undergoes conformational changes in two steps by the presence of Cu/Zn, which indicates the presence of two binding sites displaying different affinities for metals ions. Our results show that AhSOD could be classified as an intrinsically disordered protein (IDP) that is folded when metals are bound and with high thermal stability.
从皱果苋中克隆、表达并鉴定了一种新型铜锌超氧化物歧化酶。核苷酸序列分析显示一个456 bp的开放阅读框（ORF），预计编码一个分子量为15.6 kDa、pI为5.4的蛋白质。结构分析表明存在参与铜/锌结合的高度保守氨基酸残基。重组苋属超氧化物歧化酶（rAhSOD）在100℃孵育30分钟后仍具有超过50%的催化活性。对皱果苋超氧化物歧化酶（AhSOD）氨基酸序列进行的球状和无序性的计算机分析表明，该蛋白质主要是无序的；圆二色性证实了这一点，其显示缺乏二级结构。内在荧光研究表明，rAhSOD在铜/锌存在的情况下分两步发生构象变化，这表明存在两个对金属离子具有不同亲和力的结合位点。我们的结果表明，AhSOD可归类为一种内在无序蛋白质（IDP），在结合金属时折叠且具有高热稳定性。
