Qisen Zhang, Export Grains Innovation Centre, South Perth, Australia
Malt and Grains
Barley malting is a controlled seed germination process and requires several families of enzymes including α-amylase, β-amylase, α-glucosidase, limit dextrinase for starch, β-glucanase for β-glucan, and proteases for insoluble storage degradation. Barley genome sequencing and gene annotation have been completed and genomic data are available in the public domain at IPK Gatersleben. They provide useful resources for understanding malting processes on a molecular level. Gene coding for the enzymes can be extracted from the databases. Gene copy numbers, locations, and sequences can also be obtained. Next generation sequencing (NGS) technologies have also widely been used for studies of barley genetic diversities by genome resequencing and for studies of gene expression by RNA sequencing. Expression levels of functioning genes during malting may be studied by NGS technologies. Here, we have investigated genetic coding for α-amylase and proteases in the barley genome and their expression levels during malting. The barley genome contains 12 α-amylase genes (amy). They are grouped onto four subfamilies with six, three, one, and two members on subfamilies 1, 2, 3, and 4, respectively. Subfamilies 1 and 2 have high sequence identifiers within the same family and are localized on chromosomes 6 and 7, respectively. They are most important during malting, since they have expanded copy numbers. They correspond to amy1 and amy2 loci in genetic maps, which have been demonstrated to control many malting qualities. Subfamilies 1 and 2 not only have expanded gene copy numbers generated by genome duplication, but also have unique regulatory motifs on promoter regions, including motifs for GA response and gene expression enhancement. Other crops such as rice, corn, and wheat do not have the expanded gene numbers on amy families 1 and 2, nor amy gene expression enhancers on promoter regions. RNA sequencing shows that transcripts of amy subfamilies 1 and 2 are most abundant 24 hours after germination. However, transcript levels are substantially different among different malting varieties. This implicates a different starch hydrolytic rate among the varieties. Barley seeds contain a large amount of insoluble storage proteins, accounting for over 50% of total proteins with a majority being hordein. During malting, the storage proteins are mobilized by proteases. The amount of insoluble storage proteins decreases to about 40% of total proteins in malts. The barley genome contains roughly 580 protease genes. RNA-seq showed that about 180 of them are expressed during seed germination. Using LC-mass spectrometry, about 110 protease proteins are detected. Both malting barley varieties and malting conditions affect the expression levels and activities of hordein-hydrolysing proteases. A better understanding of the key proteases and their best functioning conditions during malting will make it possible to significantly improve FAN levels without over-modification.
Qisen Zhang has a Ph.D. degree in biochemistry.