Genomic, Biochemical, and Modeling Analyses of Asparagine Synthetases from Wheat

作者：Hongwei Xu, Tanya Y. Curtis, Stephen J. Powers, Sarah Raffan, Runhong Gao, Jianhua Huang, Monika Heiner, David Gilbert, Nigel G. Halford · 发表于：Frontiers in Plant Science · 年份：2018 · DOI：10.3389/fpls.2017.02237 · 被引用次数：33 · 研究领域：Potato Plant Research、Plant tissue culture and regeneration、Plant nutrient uptake and metabolism

Asparagine synthetase activity in cereals has become an important issue with the discovery that free asparagine concentration determines the potential for formation of acrylamide, a probably carcinogenic processing contaminant, in baked cereal products. Asparagine synthetase catalyses the ATP-dependent transfer of the amino group of glutamine to a molecule of aspartate to generate glutamate and asparagine. Here, asparagine synthetase-encoding polymerase chain reaction products were amplified from wheat (Triticum aestivum) cv. Spark cDNA. The encoded proteins were assigned the names TaASN1, TaASN2 and TaASN3 on the basis of comparisons with other wheat and cereal asparagine synthetases. Although very similar to each other they differed slightly in size, with molecular masses of 65.49, 65.06 and 66.24 kDa, respectively. Chromosomal positions and scaffold references were established for TaASN1, TaASN2 and TaASN3, and a fourth, more recently identified gene, TaASN4. TaASN1, TaASN2 and TaASN4 were all found to be single copy genes, located on Chromosomes 5, 3 and 4, respectively, of each genome (A, B and D), although variety Chinese Spring lacked a TaASN2 gene in the B genome. Two copies of TaASN3 were found on Chromosome 1 of each genome, and these were given the names TaASN3.1 and TaASN3.2. The TaASN1, TaASN2 and TaASN3 PCR products were heterologously-expressed in Escherichia coli (TaASN4 was not investigated in this part of the study). Western blot analysis identified two mono...