TY - JOUR
T1 - Molecular cloning and characterization of the ATP citrate lyase from carotenogenic yeast Phaffia rhodozyma
AU - Chávez-Cabrera, Cipriano
AU - Marsch, Rodolfo
AU - Bartolo-Aguilar, Yaneth
AU - Flores-Bustamante, Zoila Rosa
AU - Hidalgo-Lara, María Eugenia
AU - Martínez-Cárdenas, Anahí
AU - Cancino-Díaz, Juan Carlos
AU - Sánchez, Sergio
AU - Flores-Cotera, Luis Bernardo
N1 - Publisher Copyright:
© FEMS 2015.
PY - 2015/9
Y1 - 2015/9
N2 - ATP citrate lyase (ACL), is a key cytosolic source of acetyl-CoA for fatty acid and sterol biosynthesis and appear to be involved in carotenoid biosynthesis in yeasts. Three homologous DNA sequences encoding ACLs in Phaffia rhodozyma were isolated i.e two genes and one cDNA. The two genes were multi-intronic, with 3450-bp-coding sequences and both genes, as the cDNA, encoded identical 120.1-kDa polypeptides. Full-length amino acid sequences of these ACLs showed the two multidomains, PLN02235 and PLN02522, which are necessary for activity. The ACLs showed 82-87% similarity to putative ACLs from other basidiomycetes and 71% similarity to human ACL. The acl cDNA was used to express the heterologous ACL 6XHis-tagged which was identified using MALDI-TOF-MS. The sequenced peptides with 42.2% coverage showed 100% identity to the amino acid sequence generated in silico. The recombinant ACL purified to homogeneity showed an activity of 2 U. This is the first study to characterize a recombinant ACL from a carotenogenic yeast. The present study provides a key foundation for future studies to assess (a) the possible occurrence of alternative splicing, (b) identify the promoter(s) sequence(s) and (c) the involvement of ACL in the differential regulation of fatty acid and carotenoid biosynthesis in yeasts.
AB - ATP citrate lyase (ACL), is a key cytosolic source of acetyl-CoA for fatty acid and sterol biosynthesis and appear to be involved in carotenoid biosynthesis in yeasts. Three homologous DNA sequences encoding ACLs in Phaffia rhodozyma were isolated i.e two genes and one cDNA. The two genes were multi-intronic, with 3450-bp-coding sequences and both genes, as the cDNA, encoded identical 120.1-kDa polypeptides. Full-length amino acid sequences of these ACLs showed the two multidomains, PLN02235 and PLN02522, which are necessary for activity. The ACLs showed 82-87% similarity to putative ACLs from other basidiomycetes and 71% similarity to human ACL. The acl cDNA was used to express the heterologous ACL 6XHis-tagged which was identified using MALDI-TOF-MS. The sequenced peptides with 42.2% coverage showed 100% identity to the amino acid sequence generated in silico. The recombinant ACL purified to homogeneity showed an activity of 2 U. This is the first study to characterize a recombinant ACL from a carotenogenic yeast. The present study provides a key foundation for future studies to assess (a) the possible occurrence of alternative splicing, (b) identify the promoter(s) sequence(s) and (c) the involvement of ACL in the differential regulation of fatty acid and carotenoid biosynthesis in yeasts.
KW - Acetyl coenzyme A
KW - Astaxanthin
KW - Carotenoids
KW - Functional expression
KW - Xanthophyllomyces dendrorhous
UR - http://www.scopus.com/inward/record.url?scp=84949218264&partnerID=8YFLogxK
U2 - 10.1093/femsyr/fov054
DO - 10.1093/femsyr/fov054
M3 - Artículo
C2 - 26122278
SN - 1567-1356
VL - 15
JO - FEMS Yeast Research
JF - FEMS Yeast Research
IS - 6
ER -