PMID:24211038

Citation	Sugimori, D, Ogasawara, J, Okuda, K and Murayama, K (2014) Purification, characterization, molecular cloning, and extracellular production of a novel bacterial glycerophosphocholine cholinephosphodiesterase from Streptomyces sanglieri. J. Biosci. Bioeng. 117:422-30
Abstract	A novel metal ion-independent glycerophosphocholine cholinephosphodiesterase (GPC-CP) of Streptomyces sanglieri was purified 53-fold from culture supernatant with 1.1% recovery (583 U/mg-protein). The enzyme functions as a monomer with a molecular mass of 66 kDa. The gene encoding the enzyme consists of a 1941-bp ORF that produces a signal peptide of 38 amino acids for secretion and a 646 amino acid mature protein with a calculated molecular mass of 70,447 Da. The maximum activity was found at pH 7.2 and 40°C. The enzyme hydrolyzed glycerol-3-phosphocholine (GPC) over a broad temperature range (37-60°C) and within a narrow pH range near pH 7. The enzyme was stable at 50°C for 30 min and between pH 5-10.5. The enzyme exhibited specificity toward GPC and glycerol-3-phosphoethanolamine and hydrolyzed glycerol-3-phosphate and lysophosphatidylcholine. However, the enzyme showed no activity toward any diacylglycerophospholipids and little activity toward other glycerol-3-phosphodiesters and lysophospholipids. The enzyme was not inhibited in the presence of 2 mM SDS and Mg(2+); however, Cu(2+), Zn(2+), and Co(2+) remarkably inhibited activity. Enzyme activity was also slightly enhanced by Ca(2+), Na(+), EDTA, DTT, and 2-mercaptoethanol. During the hydrolysis of GPC at 37°C and pH 7.2, apparent Vmax and turnover number (kcat) were determined to be 24.7 μmol min(-1) mg-protein(-1) and 29.0 s(-1), respectively. The apparent Km and kcat/Km values were 1.41 mM and 20.6 mM(-1) s(-1), respectively. GPC hydrolysis by GPC-CP might represent a new metabolic pathway for acquisition of a phosphorus source in actinomycetes.
Links	PubMed Online version:10.1016/j.jbiosc.2013.10.004
Keywords
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
9ACTN:U6C6H1		GO:0047390: glycerophosphocholine cholinephosphodiesterase activity	ECO:0000314:		F		Figure 3 is showing the steady-state kinetics of GPC hydrolysis by GPC-CP.	complete CACAO 9810
9ACTN:U6C6H1	enables	GO:0047390: glycerophosphocholine cholinephosphodiesterase activity	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete

References

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