Expression, purification and preliminary crystallographic studies of alpha-amylase isozyme 1 from barley seeds.
The germinating barley seed contains two major alpha-amylase isozyme families, AMY1 and AMY2, involved in starch degradation to provide energy used by the plant embryo for growth. Many years of difficulty in growing three-dimensional crystals of natural AMY1 have now been overcome by a nonapeptide truncation of the enzyme C-terminus. The truncated enzyme was overexpressed in Pichia pastoris, purified and crystallized by the hanging-drop vapour-diffusion method using polyethylene glycol 8000 as precipitant and 2-propanol as an additive. Crystals belong to the orthorhombic space group P2(1)2(1)2, with unit-cell parameters a = 88.36, b = 72.82, c = 61.74 A and one molecule per asymmetric unit.
Robert
Xavier
X
Institut de Biologie et Chimie des Protéines, Laboratoire de Bio-Cristallographie CNRS-UMR 5086, 69367 Lyon CEDEX 07, France.
Gottschalk
Tine E
TE
Haser
Richard
R
Svensson
Birte
B
Aghajari
Nushin
N
D018360:Crystallography, X-Ray; D001467:Hordeum / Q000737:chemistry*; D007527:Isoenzymes / Q000737:chemistry; D010843:Pichia / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012639:Seeds / Q000737:chemistry; D000516:alpha-Amylases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism
D007527:Isoenzymes; D011994:Recombinant Proteins; D000516:alpha-Amylases