Tyrosine decarboxylase from Lactobacillus brevis: Soluble expression and characterization

• Published in: Protein expression and purification Vol. 94; pp. 33 - 39
• Main Authors: Zhang, Kai, Ni, Ye
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.02.2014
• Subjects: Amino Acid Sequence; Cloning, Molecular; Escherichia coli; Escherichia coli - genetics; l-DOPA; l-Tyrosine; Lactobacillus brevis; Lactobacillus brevis - enzymology; Pyridoxal-5′-phosphate (PLP); Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Soluble expression; Substrate Specificity; Tyramine; Tyrosine - metabolism; Tyrosine decarboxylase (TDC); Tyrosine Decarboxylase - biosynthesis; Tyrosine Decarboxylase - chemistry; Tyrosine Decarboxylase - genetics; Tyrosine Decarboxylase - isolation & purification; Tyramine; l-Tyrosine; Pyridoxal-5′-phosphate (PLP); Lactobacillus brevis; Tyrosine decarboxylase (TDC); Soluble expression; l-DOPA
• ISSN: 1046-5928; 1096-0279
• DOI: 10.1016/j.pep.2013.10.018

•First report on soluble expression of tdc (encoding tyrosine decarboxylase) from Lactobacillus brevis.•Glucose plays an important role in the soluble expression of rLbTDC.•rLbTDC exhibits decarboxylase activity towards l-DOPA besides l-tyrosine. Tyrosine decarboxylase (TDC, EC 4.1.1.25) is an enzyme that catalyzes the decarboxylation of l-tyrosine to produce tyramine and CO2. In this study, a 1881-bp tdc gene from Lactobacillus brevis was cloned and heterologously expressed in Escherichia coli BL21 (DE3). Glucose was discovered to play an important role in the soluble expression of rLbTDC. After optimization, recombinant TDC (rLbTDC) was achieved in excellent solubility and a yield of 224mg rLbTDC/L broth. The C-terminal His-Tagged rLbTDC was one-step purified with 90% recovery. Based on SDS–PAGE and gel filtration analysis, rLbTDC is a dimer composed of two identical subunits of approximately 70kDa. Using l-tyrosine as substrate, the specific activity of rLbTDC was determined to be 133.5U/mg in the presence of 0.2mM pyridoxal-5′-phosphate at 40°C and pH 5.0. The Km and Vmax values of rLbTDC were 0.59mM and 147.1μmolmin−1mg−1, respectively. In addition to l-tyrosine, rLbTDC also exhibited decarboxylase activity towards l-DOPA. This study has demonstrated, for the first time, the soluble expression of tdc gene from L. brevis in heterologous host.