Cloning, expression, crystallization and preliminary X-ray data analysis of norcoclaurine synthase from Thalictrum
Alessandra Pasquo1, Alessandra Bonamore, Stefano Franceschini
1ENEA Casaccia Research Centre, Dipartimento BIOTEC, Sezione Genetica e Genomica Vegetale, PO Box 2400, I-00100 Rome, Italy.
Abstract:
Norcoclaurine synthase (NCS) catalyzes the condensation of 3,4-dihydroxyphenylethylamine (dopamine) and 4-hydroxyphenylacetaldehyde (4-HPAA) as the first committed step in the biosynthesis of benzylisoquinoline alkaloids in plants. The protein was cloned, expressed and purified. Crystals were obtained at 294 K by the hanging-drop vapour-diffusion method using ammonium sulfate and sodium chloride as precipitant agents and diffract to better than 3.0 A resolution using a synchrotron-radiation source. The crystals belong to the trigonal space group P3(1)21, with unit-cell parameters a = b = 86.31, c = 118.36 A. A selenomethionine derivative was overexpressed, purified and crystallized in the same space group. A complete MAD data set was collected at 2.7 A resolution. The model is under construction.
