The DEVELOPMENT OF Β-D-Glucanases DURING THE GERMINATION OF BARLEY AND THE EFFECT OF KILNING ON INDIVIDUAL ISOENZYMES†

Abstract

Two endo-1,3;1,4-p-D-glucanase isoenzymes developed in response to gibberellic acid, during the germination of barley. Two endo-1,3-p-D-glucanases, one present in ungerminated, steeped grain, also developed but did not appear to be markedly stimulated by the hormone. A comparison of crude and partially purified malt extracts highlighted the errors that are involved in the specific determination of endo-1,3;1,4-p-glucanase activity in crude extracts. The development and effect of kilning on individual malt isoenzymes was demonstrated by carboxymethylcellulose (CM-cellulose) chromatography profiles.Kilning and dry-milling of germinated barley caused losses of 80-90% in the specific endo-1,3;1,4-p-glucanase activity. The effect was less pronounced if wet-milling was substituted for dry-milling. Extraction studies and CM-cellulose chromatography profiles indicated that both endo-1,3:1,4-pglucanase isoenzymes were heat labile and were particularly susceptible to oxidation. In contrast, endo-1,3-p-glucanase activity and cellobiase activity in malt extracts were less affected by the kilning process or extraction procedures. Preliminary results suggested that one of the endo-1,3-p-glucanase isoenzymes was more sensitive to kilning.

The (1-3, 1-4)-β-Glucanases in Malting Barley: Enzyme Survival and Genetic and Environmental Effects

“…These studies have shown that the initial activity of these enzymes is very low, but increases rapidly from the 2nd to the 4th day of the malting process. Nevertheless, the maximum values of enzyme activity ranged from the 4th, or 5th and 6th days, or even to the 11th day, in different experiments 1,3,14,20 . McFadden et al 17 verified that no mRNA encoding (1-3,1-4)-␤-glucanase is detected in ungerminated grain.…”

“…Changes in the kilning temperatures and /or duration alter these results. Brunswick et al 3 kilned the malt at 60°C for 16 h, with resulting losses from 80 to 90% in (1-3,1-4)-␤-glucanase activity. Barber et al 2 , using a malting schedule with a more progressive temperature increase (from 30 to 65°C), observed a better survival of the (1-3,1-4)-␤-glucanase activity with losses from 16 to 26%.…”

“…However, EI and EII can be readily distinguished by their apparent molecular weight, isoelectric point, carbohydrate content, and thermal stability. Enzyme I, codified by the gene Glb1 3 , has M r 28000, an isoelectric point of 8.5, and contains approximately 0.7% associated carbohydrate 26 . Enzyme II, codified by the gene Glb2 21 , has M r 33000, an isoelectric point > 10, and contains approximately 4% carbohydrate 26 .…”

Effects of Pulses of Higher Temperature on the Development of Enzyme Activity During Malting

“…Instead, ␣-amylase activity in most malting and feeding varieties is usually above the optimal level, at least when adjuncts are not required, so that differences above such a threshold have no effect on starch degradation during mashing 7 . Conversely, comparing activities of these enzymes in the kilned malt indicates a greater susceptibility of Extra enzymes to warm drying (stronger declines between 7 and 9 days), suggesting a diverse combination of differentially-thermostable isozymes was present 8 , which would justify the significance of the interaction effect (Table I). In regard to the effect of treatment, an initial higher temperature induced the higher ␤-glucanase activities, either integrated or in the green malt (Fig.…”

Transgenic barley expressing a protein-engineered, thermostable (1,3-1,4)-beta-glucanase during germination.

“…Furthermore, extraction of non-food products deposited in the endosperm of transgenic barley would be facilitated by the action of highly efficient, heat stable cell wall-degrading enzymes. The (1,3-1,4)-3-glucanases (EC 3.2.1.73) synthesized by the aleurone and scutellum tissues during barley grain germination (8) are susceptible to irreversible thermoinactivation at temperatures above 55°C (9,10), which may result in incomplete degradation of cell wall 13-glucans and limit the utility of barley for industrial processes unless a thermostable (1,3-1,4)-43-glucanase is present during extraction at elevated temperatures.…”