Engineering of L-amino acid deaminases for the production of α-keto acids from L-amino acids

Figures & data

Table 1. Comparison of α-ketoacids production using wild type and evolved L-AAD.

Produced ⍺-keto acid	Source of L-AAD	Whole-cell biocatalyst	Substrate	Wild type LAAD titer	Evolved L-AAD titer	References
Phenylpyruvic acid	L-AAD from Proteus mirabilis	Escherischia coli BL21(DE3)	L-phenylalanine	3.3 g/l with 82.2% substrate conversion rate	10 g/l with 100% substrate conversion rate	[34,35]
α-ketoisocaproate	L-AAD from Proteus vulgaris	Rhodococcus opacus DSM 43,250	L-Leucine	1,264 g/l	86,55 g/l with 94.25% conversion rate	[39–41]
	L-AAD from Proteus vulgaris	Escherischia coli BL21(DE3)	L-Leucine	69.1 g/l with 50% substrate conversion rate
α-ketoglutaric acid	L-AAD from Proteus mirabilis	Bacillus subtilis 168	L-Glutamic	12.79 g/l	89.11 g/l when Ep-PCR and gene shuffling are integrated	[46–48]
α-ketoisovaleric acid	L-AAD from Proteus myxofaciens	Escherischia coli BL21(DE3)	L-Valine	2.014 g/l	8.197 g/l	[50]
α-keto-γ-methylthiobutyric acid	L-AAD from Proteus vulgaris	Escherischia coli BL21(DE3)	L-Methionine	2.4 g/l with 71.2% bioconversion rate	increased molecular conversion rate up to 91.4%	[54]
Pyruvate	L-AAD from Proteus mirabilis	Escherischia coli BL21(DE3)	L-alanine	5.38 g/l	14.54 g/l	[59]

Figure 1. Mechanism of different enzymes catalyzing the amino acids to α-keto acids.

Figure 2. Schematic illustration of directed evolution method.

Figure 3. Diagram depicting strategies used for directed evolution in biocatalysis.

Hou Y, Hossain GS, Li J, et al. Production of phenylpyruvic acid from L-phenylalanine using an L-amino acid deaminase from Proteus mirabilis: comparison of enzymatic and whole-cell biotransformation approaches. Appl Microbiol Biotechnol. 2015;99(20):8391–8402.

 Google Scholar

Hou Y, Hossain GS, Li J, et al. Combination of phenylpyruvic acid (PPA) pathway engineering and molecular engineering of L-amino acid deaminase improves PPA production with an Escherichia coli whole-cell biocatalyst. Appl Microbiol Biotechnol. 2016;100(5):2183–2191.

 Google Scholar

Zhu Y, Li J, Liu L, et al. Production of alpha-ketoisocaproate via free-whole-cell biotransformation by Rhodococcus opacus DSM 43250 with L-leucine as the substrate. Enzyme Microb Technol. 2011;49(4):321–325.

 Google Scholar

Song Y, Li J, Shin HD, et al. Tuning the transcription and translation of L-amino acid deaminase in Escherichia coli improves alpha-ketoisocaproate production from L-leucine. PLoS One. 2017;12(6):e0179229.

 Google Scholar

Hossain GS, Li J, Shin HD, et al. Bioconversion of l-glutamic acid to alpha-ketoglutaric acid by an immobilized whole-cell biocatalyst expressing l-amino acid deaminase from Proteus mirabilis. J Biotechnol. 2014;169:112–120.

 Google Scholar

Hossain GS, Shin H, Li J, et al. Integrating error-prone PCR and DNA shuffling as an effective molecular evolution strategy for the production of α-ketoglutaric acid by l-amino acid deaminase. RSC Adv. 2016;6(52):46149–46158.

 Google Scholar

Li R, Sakir HG, Li J, et al. Rational molecular engineering of l-amino acid deaminase for production of α-ketoisovaleric acid from l-valine by Escherichia coli. RSC Adv. 2017;7(11):6615–6621.

 Google Scholar

Hossain GS, Li J, Shin HD, et al. One-step biosynthesis of alpha-keto-gamma-methylthiobutyric acid from L-methionine by an Escherichia coli whole-cell biocatalyst expressing an engineered L-amino acid deaminase from Proteus vulgaris. PLoS One. 2014;9(12):e114291.

 Google Scholar

Hossain GS, Shin H, Li J, et al. Transporter engineering and enzyme evolution for pyruvate production from D/L-alanine with a whole- cell biocatalyst expressing L-amino acid deaminase from Proteus mirabilis. RSC Adv. 2016;6(2016):82676–82684.

 Google Scholar