TY - JOUR
T1 - Retinal biosynthesis in fungi
T2 - Characterization of the carotenoid oxygenase CarX from Fusarium fujikuroi
AU - Prado-Cabrero, Alfonso
AU - Scherzinger, Daniel
AU - Avalos, Javier
AU - Al-Babili, Salim
PY - 2007/4
Y1 - 2007/4
N2 - The car gene cluster of the ascomycete Fusarium fujikuroi encodes two enzymes responsible for torulene biosynthesis (CarRA and CarB), an opsin-like protein (CarO), and a putative carotenoid cleaving enzyme (CarX). It was presumed that CarX catalyzes the formation of the major carotenoid in F. fujikuroi, neurosporaxanthin, a cleavage product of torulene. However, targeted deletion of carX did not impede neurosporaxanthin biosynthesis. On the contrary, ΔcarX mutants showed a significant increase in the total carotenoid content, indicating an involvement of CarX in the regulation of the pathway. In this work, we investigated the enzymatic activity of CarX. The expression of the enzyme in β-carotene-accumulating Escherichia coli cells led to the formation of the opsin chromophore retinal. The identity of the product was proven by high-performance liquid chromatography and gas chromatography-mass spectrometry. Subsequent in vitro assays with heterologously expressed and purified CarX confirmed its β-carotene-cleaving activity and revealed its capability to produce retinal also from other substrates, such as γ-carotene, torulene, and β-apo-8′-carotenal. Our data indicate that the occurrence of at least one β-ionone ring in the substrate is required for the cleavage reaction and that the cleavage site is determined by the distance to the β-ionone ring. CarX represents the first retinal-synthesizing enzyme reported in the fungal kingdom so far. It seems likely that the formed retinal is involved in the regulation of the carotenoid biosynthetic pathway via a negative feedback mechanism.
AB - The car gene cluster of the ascomycete Fusarium fujikuroi encodes two enzymes responsible for torulene biosynthesis (CarRA and CarB), an opsin-like protein (CarO), and a putative carotenoid cleaving enzyme (CarX). It was presumed that CarX catalyzes the formation of the major carotenoid in F. fujikuroi, neurosporaxanthin, a cleavage product of torulene. However, targeted deletion of carX did not impede neurosporaxanthin biosynthesis. On the contrary, ΔcarX mutants showed a significant increase in the total carotenoid content, indicating an involvement of CarX in the regulation of the pathway. In this work, we investigated the enzymatic activity of CarX. The expression of the enzyme in β-carotene-accumulating Escherichia coli cells led to the formation of the opsin chromophore retinal. The identity of the product was proven by high-performance liquid chromatography and gas chromatography-mass spectrometry. Subsequent in vitro assays with heterologously expressed and purified CarX confirmed its β-carotene-cleaving activity and revealed its capability to produce retinal also from other substrates, such as γ-carotene, torulene, and β-apo-8′-carotenal. Our data indicate that the occurrence of at least one β-ionone ring in the substrate is required for the cleavage reaction and that the cleavage site is determined by the distance to the β-ionone ring. CarX represents the first retinal-synthesizing enzyme reported in the fungal kingdom so far. It seems likely that the formed retinal is involved in the regulation of the carotenoid biosynthetic pathway via a negative feedback mechanism.
UR - http://www.scopus.com/inward/record.url?scp=34247874569&partnerID=8YFLogxK
U2 - 10.1128/EC.00392-06
DO - 10.1128/EC.00392-06
M3 - Article
C2 - 17293483
AN - SCOPUS:34247874569
SN - 1535-9778
VL - 6
SP - 650
EP - 657
JO - Eukaryotic Cell
JF - Eukaryotic Cell
IS - 4
ER -