4-Hydroxyphenylpyruvic acid (4-HPPA)

a kind of α-keto acid

is an intermediate in the metabolism of tyrosine and has a wide range of application in food

pharmaceutical and chemical industry. Using amino acids as raw material to produce the corresponding α-keto acid is thought to be both economic and efficient. Among the enzymes that convert amino acid to α-keto acid

membrane bound L-amino acid deaminase (mL-AAD)

which is anchored to the outer side of the cytomembrane

becomes an ideal enzyme to prepare α-keto acid since there is no cofactors needed and H

2

O

2

production during the reaction. In this study

the mL-AAD from Proteus vulgaris was used to prepare whole-cell catalysts to produce 4-HPPA from L-tyrosine. The secretory efficiency of mL-AAD conducted by its own twin-arginine signal peptide (twin-arginine translocation pathway

Tat) and integrated pelB (the general secretory pathway

Sec)-Tat signal peptide was determined and compared firstly

using two pET systems (pET28a and pET20b). It was found that the Tat pathway (pET28a-mlaad) resulted in higher cell-associated mL-AAD activity and cell biomass

and was more b

eneficial to prepare biocatalyst. In addition

expression hosts Bl21(DE3) and 0.05 mmol·L

-1

IPTG were found to be suitable for mL-AAD expression. The reaction conditions for mL-AAD were optimized and 72.72 mmol·L

-1

4-HPPA was obtained from 100 mmol·L

-1

tyrosine in 10 h under the optimized conditions. This bioprocess

which is more eco-friendly and economical than the traditional chemical synthesis ways

has great potential for industrial application.