Srisimarat Wiraya, Murakami Shuichiro, Pongsawasdi Piamsook, Krusong Kuakarun
Starch and Cyclodextrin Research Unit, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Phyathai Road, Bangkok 10330, Thailand.
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2013 Sep;69(Pt 9):1004-6. doi: 10.1107/S1744309113020319. Epub 2013 Aug 19.
Amylomaltase (AM; EC 2.4.1.25) belongs to the 4-α-glucanotransferase group of the α-amylase family. The enzyme can produce cycloamylose or large-ring cyclodextrin through intramolecular transglycosylation or cyclization reactions of α-1,4-glucan. Amylomaltase from the mesophilic bacterium Corynebacterium glutamicum (CgAM) contains extra residues at the N-terminus for which the three-dimensional structure is not yet known. In this study, CgAM was overexpressed and purified to homogeneity using DEAE FF and Phenyl FF columns. The purified CgAM was crystallized by the vapour-diffusion method. Preliminary X-ray data showed that the CgAM crystal diffracted to 1.7 Å resolution and belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 73.28, b = 82.61, c = 118.64 Å. To obtain the initial phases, crystals of selenomethionyl-substituted amylomaltase were produced, and multiple-wavelength anomalous dispersion phasing and structure refinement are now in progress.
淀粉麦芽糖酶（AM；EC 2.4.1.25）属于α-淀粉酶家族的4-α-葡聚糖转移酶组。该酶可通过α-1,4-葡聚糖的分子内转糖基化或环化反应产生环糊精或大环环糊精。来自嗜温细菌谷氨酸棒杆菌的淀粉麦芽糖酶（CgAM）在N端含有额外的残基，其三维结构尚不清楚。在本研究中，使用DEAE FF和苯基FF柱对CgAM进行了过表达和纯化，使其达到均一性。通过气相扩散法使纯化的CgAM结晶。初步X射线数据显示，CgAM晶体的衍射分辨率为1.7 Å，属于空间群P2(1)2(1)2(1)，晶胞参数a = 73.28、b = 82.61、c = 118.64 Å。为了获得初始相位，制备了硒代甲硫氨酸取代的淀粉麦芽糖酶晶体，目前正在进行多波长反常色散相位分析和结构精修。
