Cloning, expression and characterization of phenylalanine ammonia-lyase from Rhodotorula glutinis

Zhu, Liping; Cui, Wenjing; Fang, Yueqin; Liu, Yi; Gao, Xinxing; Zhou, Zhemin

doi:10.1007/s10529-013-1140-7

Cited by 49 publications

(52 citation statements)

References 18 publications

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“…However, k cat /K m of RgPAL/TAL was significantly greater for L-phenylalanine than for L-tyrosine. This result is consistent with previous research (23,29). However, the main strategy of synthetic bioengineering for phenylpropanoid biosynthesis is to use p-coumaric acid as the initial substrate rather than transcinnamic acid.…”

Section: )supporting

confidence: 93%

A novel process for obtaining phenylpropanoic acid precursor using Escherichia coli with a constitutive expression system

Liang

Guo

Jiang

et al. 2016

Food Sci Biotechnol

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Phenylpropanoids are widely used in food supplements, pharmaceuticals, and cosmetics with diverse benefits to human health. -cinnamic acid or-coumaric acid is usually used as the starting precursor to produce phenylpropanoids. Synthetic bioengineering of microbial cell factories offers a sustainable and flexible alternative method for obtaining these compounds. In this study, a constitutive expression system consisting of phenylalanine/tyrosine ammonia lyase was developed to produce a phenylpropanoic acid precursor in. To improve -cinnamic acid and-coumaric acid production, BioBrick optimization was investigated, causing a 7.2- and 14.2-fold increase in the yield of these compounds, respectively. The optimum strain was capable of producing 78.81 mg/L of-cinnamic acid and 34.67 mg/L of -coumaric acid in a shake flask culture. The work presented here paves the way for the development of a sustainable and economical process for microbial production of a phenylpropanoic acid precursor.

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Section: )supporting

confidence: 93%

A novel process for obtaining phenylpropanoic acid precursor using Escherichia coli with a constitutive expression system

Liang

Guo

Jiang

et al. 2016

Food Sci Biotechnol

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“…Thus, the optimal temperature of ZmPAL2 was 55°C. It was higher than that in R. chinensis and R. glutinis [17,23], similar to the ZmPAL1 and CdPAL from C. deserticola [29], and lower than that in S. miltiorrhiza [14]. As for pH, the pH profiles for the purified ZmPAL2 were shown in Fig.…”

Section: Biochemical Property Analysismentioning

confidence: 67%

“…It was consistent with the result of sequence analysis. Among those reported PALs, the recombinant ZmPAL2 exhibited a high level of PAL activity, which was higher than BoPAL1 (2.26 U/mg) and BoPAL2 (3.77 U/mg) from B. oldhamii and RgPAL (4.3 U/mg) from R. glutinis [18,39,23]. Among the reported PALs, ZmPAL2 exhibited a high level of PAL activity.…”

Section: Expression and Purification Of The Recombinant Zmpal2mentioning

confidence: 95%

Molecular Characterization of a Recombinant Zea mays Phenylalanine Ammonia-Lyase (ZmPAL2) and Its Application in trans-Cinnamic Acid Production from l-Phenylalanine

Zang

Jiang

Cong

et al. 2015

Appl Biochem Biotechnol

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Phenylalanine ammonia-lyase (PAL) is one of the most extensively studied enzymes with its crucial role in secondary phenylpropanoid metabolism of plants. Recently, its demand has been increased for aromatic chemical production, but its applications in trans-cinnamic acid production were not much explored. In the present study, a putative PAL gene from Zea mays designated as ZmPAL2 was expressed and characterized in Escherichia coli BL21 (DE3). The recombinant ZmPAL2 exhibited a high PAL activity (7.14 U/mg) and a weak tyrosine ammonia-lyase activity. The optimal temperature of ZmPAL2 was 55 °C, and the thermal stability results showed that about 50 % of enzyme activity remained after a treatment at 60 °C for 6 h. The recombinant ZmPAL2 is a good candidate for the production of trans-cinnamic acid. The vitro conversion indicated that the recombinant ZmPAL2 could effectively catalyze the L-phenylalanine to trans-cinnamic acid, and the trans-cinnamic acid concentration can reach up to 5 g/l.

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“…Several enzymes have previously been characterized both in vivo and in vitro, and the data are available from both patent and scientific literature (3,(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32). However, the conditions of kinetic analysis are often quite different and may not represent the extent to which the enzymes perform in vivo.…”

mentioning

confidence: 99%

“…Enzymes categorized as acting on either of the structurally similar amino acids tyrosine or phenylalanine often have some activity for the other substrate as well (16,32) or for similar compounds, such as 3,4-dihydroxy-L-phenylalanine (L-dopa) (22). Homologous and structurally similar enzymes are tyrosine 2,3-aminomutases (TAM; EC 5.4.3.6) and phenylalanine aminomutase (PAM; EC 5.4.3.11) (33)(34)(35).…”

mentioning

confidence: 99%

Highly Active and Specific Tyrosine Ammonia-Lyases from Diverse Origins Enable Enhanced Production of Aromatic Compounds in Bacteria and Saccharomyces cerevisiae

Jendresen

Stahlhut

et al. 2015

Appl Environ Microbiol

174

183

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Phenylalanine and tyrosine ammonia-lyases form cinnamic acid and p-coumaric acid, which are precursors of a wide range of aromatic compounds of biotechnological interest. Lack of highly active and specific tyrosine ammonia-lyases has previously been a limitation in metabolic engineering approaches. We therefore identified 22 sequences in silico using synteny information and aiming for sequence divergence. We performed a comparative in vivo study, expressing the genes intracellularly in bacteria and yeast. When produced heterologously, some enzymes resulted in significantly higher production of p-coumaric acid in several different industrially important production organisms. Three novel enzymes were found to have activity exclusively for phenylalanine, including an enzyme from the low-GC Gram-positive bacterium Brevibacillus laterosporus, a bacterial-type enzyme from the amoeba Dictyostelium discoideum, and a phenylalanine ammonia-lyase from the moss Physcomitrella patens (producing 230 M cinnamic acid per unit of optical density at 600 nm [OD 600 ]) in the medium using Escherichia coli as the heterologous host). Novel tyrosine ammonia-lyases having higher reported substrate specificity than previously characterized enzymes were also identified. Enzymes from Herpetosiphon aurantiacus and Flavobacterium johnsoniae resulted in high production of p-coumaric acid in Escherichia coli (producing 440 M p-coumaric acid OD 600 unit ؊1 in the medium) and in Lactococcus lactis. The enzymes were also efficient in Saccharomyces cerevisiae, where p-coumaric acid accumulation was improved 5-fold over that in strains expressing previously characterized tyrosine ammonia-lyases. Small organic molecules of biotechnological interest include aromatic structures that are derived from p-coumaric acid (pHCA). pHCA can be formed from phenylalanine either through deamination to cinnamic acid (CA) by phenylalanine ammonialyase (PAL; EC 4.3.1.24) and subsequent hydroxylase activity by trans-cinnamate-4-monooxygenase or through deamination of tyrosine by tyrosine ammonia-lyase (TAL; EC 4.3.1.23) (Fig. 1). Considering that the route from phenylalanine requires activity of a P450 enzyme, which has been shown to be the limiting step in previous engineering strategies (1, 2), deamination of tyrosine for production of pHCA may be preferred in microbial cell factories. Tyrosine ammonia-lyases (TAL) have been employed for the production of plant biochemicals and aromatic compounds, e.g., for pHCA itself (3-5), or in the production of stilbenes, such as resveratrol (6, 7), flavonoids, such as naringenin (8, 9), cinnamoyl anthranilates (10), or plastic precursors, such as p-hydroxystyrene (11, 12), yet the TAL reaction may be the limiting step (10, 13).Due to the significant interest in production of phenolic compounds in biotechnological processes, we aimed at increasing the range of characterized phenylalanine ammonia-lyases (PAL) and tyrosine ammonia-lyases (TAL), and in particular at identifying the optimal TAL for use in microbial cell factories...

show abstract

Cloning, expression and characterization of phenylalanine ammonia-lyase from Rhodotorula glutinis

Cited by 49 publications

References 18 publications

A novel process for obtaining phenylpropanoic acid precursor using Escherichia coli with a constitutive expression system

A novel process for obtaining phenylpropanoic acid precursor using Escherichia coli with a constitutive expression system

Molecular Characterization of a Recombinant Zea mays Phenylalanine Ammonia-Lyase (ZmPAL2) and Its Application in trans-Cinnamic Acid Production from l-Phenylalanine

Highly Active and Specific Tyrosine Ammonia-Lyases from Diverse Origins Enable Enhanced Production of Aromatic Compounds in Bacteria and Saccharomyces cerevisiae

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