1. Overproduction, purification and preliminary X-ray diffraction analysis of a sulfotransferase from Mycobacterium tuberculosis H37Rv
Shotaro Tanaka, Yuuji Moriizumi, Makoto Kimura, Yoshimitsu Kakuta Acta Crystallogr Sect F Struct Biol Cryst Commun. 2005 Jan 1;61(Pt 1):33-5. doi: 10.1107/S1744309104022328. Epub 2004 Sep 25.
Sulfotransferase STF1 from the Mycobacterium tuberculosis H37Rv genome was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. The crystals diffract to 1.5 A resolution using synchrotron radiation at SPring-8. The crystals are monoclinic and belong to space group P2(1), with unit-cell parameters a = 40.86, b = 95.76, c = 48.04 A, beta = 106.43 degrees. The calculated Matthews coefficient is approximately 2.1 A3 Da(-1) assuming the presence of one molecule of STF1 in the asymmetric unit. A substrate-binding assay using a PAP-agarose column suggests that STF1 exhibits sulfotransferase activity.
2. Crystallization and preliminary X-ray analysis of native and selenomethionine 2-hydroxybiphenyl 3-monooxygenase
Andreas Meyer, Daniel Tanner, Andreas Schmid, David F Sargent, Hans-Peter E Kohler, Bernard Witholt Acta Crystallogr D Biol Crystallogr. 2003 Apr;59(Pt 4):741-3. doi: 10.1107/s0907444903002634. Epub 2003 Mar 25.
2-hydroxybiphenyl 3-monooxygenase (HbpA; EC 1.14.13.44) from Pseudomonas azelaica HBP1 was produced in Escherichia coli both as native and SeMet-labelled protein. The two enzymes were purified to homogeneity and crystallized by the hanging-drop vapour-diffusion method. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 108.6, b = 196.8, c = 79.3 A, beta = 97.7 degrees for the native protein and a = 108.3, b = 196.8, c = 79.0 A, beta = 97.8 degrees for SeMet HbpA. Crystal-packing considerations led to the assumption of two HbpA subunits per asymmetric unit, which corresponds to a V(M) value of 3.3 A(3) Da(-1) and a solvent content of 62%. The crystals were radiation-sensitive and only had a lifespan of about 120 s when exposed to synchrotron radiation on an undulator beamline. To obtain complete data sets, data were collected from 23 native and 26 derivative crystals. The high-resolution limit was 2.0 A for native and 2.25 A for SeMet HbpA.
3. Crystallization and preliminary crystallographic analysis of human glycosylated haemoglobin
Vitaly E Syakhovich, N T Saraswathi, Marc Ruff, Sergey B Bokut, Dino Moras Acta Crystallogr Sect F Struct Biol Cryst Commun. 2006 Feb 1;62(Pt 2):106-9. doi: 10.1107/S1744309105042764. Epub 2006 Jan 27.
Human glycosylated haemoglobin A1C is a stable minor variant formed in vivo by post-translational modification of the main form of haemoglobin by glucose. Crystals of oxyHbA1C were obtained using the hanging-drop vapour-diffusion method and PEG as precipitant. The diffraction pattern of the crystal extends to a resolution of 2.3 A at 120 K. The crystals belong to space group C2, with unit-cell parameters a = 237.98, b = 59.27, c = 137.02 A, alpha = 90.00, beta = 125.40, gamma = 90.00 degrees. The presence of two and a half molecules per asymmetric unit gives a crystal volume per protein weight (VM) of 9.70 A3 Da(-1) and a solvent content of 49%.
