Rhodotorula gracilis

D-Aminoacid oxidase has been isolated from a nnmber of yeasts, and the nucleotide sequence of the enzyme from Rhodotorula gracilis ATCC 26217 has been established (Alonso et al. 1998). The gene could be overexpressed in Escherichia coli, and levels of the enzyme were greater under conditions of low aeration the enzyme isolated from the recombinant organisms was apparently the apoenzyme since maximum activity required the presence of FAD. 

Alonso J, JL Barredo, B Diez, E Mellado, E Salto, JL Garcia, E Cortes (1998) D-amino-acid oxidase gene from Rhodotorula gracilis (Rhodosporidium toruloides) ATCC 26217. Microbiology (UK) 144 1095-1101. 

The use of D-AAO from the yeast Rhodotorula gracilis to deracemize naphthyl amino acids has been studied in some detail by the groups of Servi and Pollegioni, who compared the kinetic properties of the enzyme with racemic 1- and 2-naphthylalanine (1 and 2) and 1- and 2-naphthylglycine (3 and 4). 

DAAO is one of the most extensively studied flavoprotein oxidases. The homodimeric enzyme catalyzes the strictly stere-ospecihc oxidative deamination of neutral and hydrophobic D-amino acids to give a-keto acids and ammonia (Fig. 3a). In the reductive half-reaction the D-amino acid substrate is converted to the imino acid product via hydride transfer (21). During the oxidative half-reaction, the imino acid is released and hydrolyzed. Mammalian and yeast DAAO share the same catalytic mechanism, but they differ in kinetic mechanism, catalytic efficiency, substrate specificity, and protein stability. The dimeric structures of the mammalian enzymes show a head-to-head mode of monomer-monomer interaction, which is different from the head-to-tail mode of dimerization observed in Rhodotorula gracilis DAAO (20). Benzoate is a potent competitive inhibitor of mammalian DAAO. Binding of this ligand strengthens the apoenzyme-flavin interaction and increases the conformational stability of the porcine enzyme. 

Figure 5 Kinetics of transport of D- and L-xylose in the yeast Rhodotorula gracilis. [Adapted from Dusinsky (I).]...

The first DAAO studied mechanistically was from pig kidney (pkDAAO) many kinetic and mechanistic studies have been performed on this enzyme. More recently, yeast DAAOs from Rhodotorula gracilis (RgDAAO) and Trigonopsis variabilis (TvDAAO) have also been studied. Each has different substrate specificities. The best substrate for pkDAAO is D-proline, followed by hydrophobic and neutral amino acids. Positively charged amino acids are bad substrates, while negatively charged D-amino acids are not substrates.In contrast, methionine and valine are the best substrates for RgDAAO. ... 

Biological Rhodotorula gracilis, a yeast isolated from an insecticide-treated soil, degraded chlorobenzilate in a basal medium supplemented by sucrose. Metabolites identified by this decarboxylation process were 4,4 -dichlorobenzilic acid, 4,4 -dichloroben-zophenone and carbon dioxide (Miyazaki et al., 1969, 1970). 

Kessell, R.H.J. (1968) Fatty adds of Rhodotorula gracilis fat production of submerged culture and the particular effect of pH value. /. Appl. Bactenol. 31, 220-231. 

Shashi, K., Bachhawat, A.K. and Joseph, R. (1990) ATP citrate lyase of Rhodotorula gracilis purification and properties. Biochim. Biophys. Acta 1033, 23-30. 

Phenylalanine ammonialyase EC 4.3.1.5 Rhodotorula gracilis Cinnamic acid (NHi -salt) L-Phenylalanine... 

However, the en2ymes of both pathways have been demonstrated in Candida utilis where xylose isomerase was found inducible and isomerization pathway occurred when xylose as the carbon source was alone in the medium [94, 95]. When both pentoses and hexoses were supplied to the medium, the conversion was carried out through oxido-reductive pathway. Indirect evidence for the presence of xylose isomerase in Rhodotorula gracilis and other yeast hke organisms has also been reported [96, 97]. 

CONTROL OF ACYL LIPID DESATURATION IN RHODOTORULA GRACILIS... 

When grown under conditions of nitrogen-limitation, the carotenoid pigmented yeast Rhodotorula gracilis has been shown to accumulate large quantities of triacylglycerols. However, to date, the commercial utilization of this storage lipid has been limited by its unsaturated nature. 

THE EFFECTS OF PACLOBUTRAZOL ON STEROL AND ACYL LIPID COMPOSITION OF MEMBRANES IN APIUM GRAVEOLENS AND RHODOTORULA GRACILIS... 

Apium graveolens and Rhodotorula gracilis were grown (see [6] [9]) in the presence or absence of paclobutrazol (50 and 75 iM respectively). Plant and yeast cells were harvested after 7 days and 2 days growth, respectively. Membranes were isolated (see [10] [11]) and sterol and phospholipid compositions determined (see [6] [9]) Membrane fluidity was measured by fluorescence polarization using DPH as a probe [12]. 

Acyl Lipid Metabolism in Rhodotorula gracilis (CBS 3043) and the Effects of Methyl Sterculate on Fatty Acid... 

ACYL LIPID METABOLISM IN RHODOTORULA GRACILIS (CBS 3043) AND THE EFFECTS OF METHYL STERCULATE ON FATTY ACID DESATURATION... 

C.E. Rolph, R.S. Moreton, I.S. Small and J.L. Harwood, Acyl lipid metabolism and fatty acid desaturation in the yeast Rhodotorula gracilis (CBS 3043), Blochem.Soc.Trans, (in press). 

In a similar dynamic kinetic resolution method, the DAAO from Rhodotorula gracilis was recently combined in a one-pot reaction with i-aspartate amino transferase (l-AAT) from Escherichia coli and used for the deracemization of racemic 2-naphthylalanine (DL-2-NAla) (Scheme 11.13b). After the oxidation of the D-amino acid catalyzed by DAAO, the formed 2-naphthyl pyruvate (2-NPA) served as a substrate of l-AAT in the presence of cysteine sulfinic acid (CSA) as an amino donor. Almost quantitative yields of the enantiomerically pure t-2-NAla were achieved by using this cascade reaction, thanks to the spontaneous degradation of the 3-keto sulfinic acid produced in the t-AAT-catalyzed transamination that drives the overall reaction equilibrium toward the desired direction [26]. 

Hbfer, M., Betz, A., Kotyk, A., 1971. Metabolism of the obligatory aerobic yeast Rhodotorula gracilis IV. Induction of an enzyme necessary for d-xylose catabolism. Biochimica et Biophysica Acta -General Subjects 252 (1), 1-12. 

Naphthylamine D-Amino acid oxidase Rhodotorula gracilis), L-aspartate aminotransferase Esch ichia coli) (rflc)-2-Naphtylamine + cysteine sulfinic acid DE [50]... 

For the preparation of 2-naphthyl alanine, a one-pot, two-step enzyme cascade was invented with d-AAO from Rhodotorula gracilis and an L-aminotransferase from E. coli [50]. Detail information about the deracemization steps was provided in Section 29.3.1 and shown in Scheme 29.3. Starting with the racemic 2-naphthyl alanine, the AAO generated the corresponding a-keto acid, which served as rfie substrate in the reductive amination step to enantiopure 2-naphthyl alanine. An irreversible amino donor, cysteine sulfinic acid, was used. 

Choi, S.Y., Ryu, D.D.Y., Rhee, J.S., 1982. Production of microbial lipid effects of growth rate and oxygen on lipid synthesis and fatty acid composition of Rhodotorula gracilis. Biotechnology and Bioengineering 24, 1165—1172. 

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