Abstract
Cysteine sulfinic acid decarboxylase catalyzes the last step of taurine biosynthesis in mammals, and belongs to the fold type I superfamily of pyridoxal-5′-phosphate (PLP)-dependent enzymes. Taurine (2-aminoethanesulfonic acid) is the most abundant free amino acid in animal tissues; it is highly present in liver, kidney, muscle, and brain, and plays numerous biological and physiological roles. Despite the importance of taurine in human health, human cysteine sulfinic acid decarboxylase has been poorly characterized at the biochemical level, although its three-dimensional structure has been solved. In the present work, we have recombinantly expressed and purified human cysteine sulfinic acid decarboxylase, and applied a simple spectroscopic direct method based on circular dichroism to measure its enzymatic activity. This method gives a significant advantage in terms of simplicity and reduction of execution time with respect to previously used assays, and will facilitate future studies on the catalytic mechanism of the enzyme. We determined the kinetic constants using L-cysteine sulfinic acid as substrate, and also showed that human cysteine sulfinic acid decarboxylase is capable to catalyze the decarboxylation—besides its natural substrates L-cysteine sulfinic acid and L-cysteic acid—of L-aspartate and L-glutamate, although with much lower efficiency.
Highlights
IntroductionMany works have shown that taurine plays numerous biological and physiological roles [2]
Human cysteine sulfinic acid decarboxylase (CSAD) (EC 4.1.1.29), whose cDNA was cloned in pNIC28-Bsa4, was expressed in the E. coli Rosetta(DE3) strain with an N-terminal tag of 23 residues including a hexahistidine stretch
Most of the expressed protein was found in the inclusion bodies, the purification yield was of about 3 mg of soluble protein per liter of bacterial culture
Summary
Many works have shown that taurine plays numerous biological and physiological roles [2]. Does it conjugate bile acids [3], but it plays a role as antioxidant [4,5], osmoregulator [6], and membrane stabilizer [7]. In the central nervous system, taurine acts as neurotransmitter, neuroprotective agent, and regulator of intracellular calcium homeostasis [8]. Low levels of taurine are associated with various pathological conditions, including cardiomyopathy [9], retinal degeneration [10], prenatal and postnatal growth retardation [11], and obesity [12]
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