TY - JOUR
T1 - Molecular and functional characterization of an invertase secreted by Ashbya gossypii
AU - Aguiar, Tatiana Q.
AU - Dinis, Cláudia
AU - Magalhães, Frederico
AU - Oliveira, Carla
AU - Wiebe, Marilyn G.
AU - Penttilä, Merja
AU - Domingues, Lucília
N1 - CA2: BA2131
CA2: BA311
SDA: BIC
ISI: BIOCHEMISTRY & MOLECULAR BIOLOGY
PY - 2014/1/1
Y1 - 2014/1/1
N2 - The repertoire of hydrolytic enzymes natively secreted by the filamentous fungus Ashbya (Eremothecium) gossypii has been poorly explored. Here, an invertase secreted by this flavinogenic fungus was for the first time molecularly and functionally characterized. Invertase activity was detected in A. gossypii culture supernatants and cell-associated fractions. Extracellular invertase migrated in a native polyacrylamide gel as diffuse protein bands, indicating the occurrence of at least two invertase isoforms. Hydrolytic activity toward sucrose was approximately 10 times higher than toward raffinose. Inulin and levan were not hydrolyzed. Production of invertase by A. gossypii was repressed by the presence of glucose in the culture medium. The A. gossypii invertase was demonstrated to be encoded by the AFR529W (AgSUC2) gene, which is highly homologous to the Saccharomyces cerevisiae SUC2 (ScSUC2) gene. Agsuc2 null mutants were unable to hydrolyze sucrose, proving that invertase is encoded by a single gene in A. gossypii. This mutation was functionally complemented by the ScSUC2 and AgSUC2 genes, when expressed from a 2-μm-plasmid. The signal sequences of both AgSuc2p and ScSuc2p were able to direct the secretion of invertase into the culture medium in A. gossypii.
AB - The repertoire of hydrolytic enzymes natively secreted by the filamentous fungus Ashbya (Eremothecium) gossypii has been poorly explored. Here, an invertase secreted by this flavinogenic fungus was for the first time molecularly and functionally characterized. Invertase activity was detected in A. gossypii culture supernatants and cell-associated fractions. Extracellular invertase migrated in a native polyacrylamide gel as diffuse protein bands, indicating the occurrence of at least two invertase isoforms. Hydrolytic activity toward sucrose was approximately 10 times higher than toward raffinose. Inulin and levan were not hydrolyzed. Production of invertase by A. gossypii was repressed by the presence of glucose in the culture medium. The A. gossypii invertase was demonstrated to be encoded by the AFR529W (AgSUC2) gene, which is highly homologous to the Saccharomyces cerevisiae SUC2 (ScSUC2) gene. Agsuc2 null mutants were unable to hydrolyze sucrose, proving that invertase is encoded by a single gene in A. gossypii. This mutation was functionally complemented by the ScSUC2 and AgSUC2 genes, when expressed from a 2-μm-plasmid. The signal sequences of both AgSuc2p and ScSuc2p were able to direct the secretion of invertase into the culture medium in A. gossypii.
KW - Ashbya gossypii
KW - Glucose repression
KW - Invertase secretion
KW - Secretion regulation
UR - http://www.scopus.com/inward/record.url?scp=84900813550&partnerID=8YFLogxK
U2 - 10.1007/s12033-013-9726-9
DO - 10.1007/s12033-013-9726-9
M3 - Article
C2 - 24452331
AN - SCOPUS:84900813550
SN - 1073-6085
VL - 56
SP - 524
EP - 534
JO - Molecular Biotechnology
JF - Molecular Biotechnology
IS - 6
ER -