Lysine and threonine biosynthesis in sorghum seeds: characterisation of aspartate kinase and homoserine dehydrogenase isoenzymes

Autor(es)

Ferreira, R. R. ^[1] ; Meinhardt, L. W. ; Azevedo, R. A. ^[3]

Afiliação do(s) autor(es)

^[1] Universidade de São Paulo (USP). Escola Superior de Agricultura Luiz de Queiroz. - Brasil

^[3] Universidade de São Paulo (USP). Escola Superior de Agricultura Luiz de Queiroz. - Brasil

• Tipo de documento: Artigo Científico
• Fonte: Annals of Applied Biology; v. 149, n. 1, p. 77-86, Aug. 2006.
• Área do conhecimento: Ciências Biológicas - Botânica
• Assunto(s): Sementes Cereais Sorgo Aminoácidos Lisina Treonina Aspartato quinase

Resumo

Aspartate kinase (AK, EC 2.7.2.4) and homoserine dehydrogenase (HSDH, EC 1.1.1.3) have been partially purified and characterised from immature sorghum seeds. Two peaks of AK activity were eluted by anion-exchange chromatography [diethylaminoethyl (DEAE)-Sephacel] with 183 and 262 mM KCl, and both activities were inhibited by lysine. Similarly, two peaks of HSDH activity were eluted with 145 and 183 mM KCl; the enzyme activity in the first peak in elution order was shown to be resistant to threonine inhibition, whereas the second was sensitive to threonine inhibition. However, following gel filtration chromatography (Sephacryl S-200), one peak of AK activity co-eluted with HSDH and both activities were sensitive to threonine inhibition, suggesting the presence of a bifunctional threonine-sensitive AK–HSDH isoenzyme with a molecular mass estimated as 167 kDa. The activities of AK and HSDH were studied in the presence of lysine, threonine, methionine, valine, calcium, ethylene glycol bis(2-aminoethylether)-N,N,N'N'-tetraacetic acid, calmodulin, S-adenosylmethionine (SAM), S-2-aminoethyl-l-cysteine (AEC) and increasing concentrations of KCl. AK was shown to be inhibited by threonine and lysine, confirming the existence of two isoenzymes, one sensitive to threonine and the other sensitive to lysine, the latter being predominant in sorghum seeds. Methionine, SAM plus lysine and AEC also inhibited AK activity; however, increasing KCl concentrations and calcium did not produce any significant effect on AK activity, indicating that calcium does not play a role in AK regulation in sorghum seeds. HSDH also exhibited some inhibition by threonine, but the majority of the activity was not inhibited, thus indicating the existence of a threonine-sensitive isoenzyme and a second predominant threonine-insensitive isoenzyme. Valine and SAM plus threonine also inhibited HSDH; however, increasing concentrations of KCl and calcium had no inhibitory effect. (AU)

• Processo FAPESP: 00/05960-2 - Isolamento, purificacao e caracterizacao bioquimica e genetica de enzimas da via de biossintese de lisina em sorgo (sorghum ssp).
• Beneficiário: Renato Rodrigues Ferreira
• Modalidade de apoio: Bolsas no Brasil - Doutorado
• Processo FAPESP: 01/13216-4 - Estudo do metabolismo de lisina em sorgo.
• Beneficiário: Ricardo Antunes de Azevedo
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 04/16039-4 - Entendendo o metabolismo de lisina em cereais
• Beneficiário: Ricardo Antunes de Azevedo
• Modalidade de apoio: Auxílio à Pesquisa - Temático