A second project designed to improve the general efficiency of feed utilization in sheep, and to increase specifically nutrient supply to sheep wool follicles involves the introduction of the glyoxylate cycle to sheep. The rationale for this research stems from the fact that the microorganisms that populate the rumen in sheep consume essentially all available carbohydrate in the ingested feed and produce a range of fermentation products, the most important of which, from an energy viewpoint, are the volatile fatty acids. After absorption by the sheep, these are used directly for energy or, if gluconeogenic, are converted to glucose to provide the carbohydrate that is essential for the proper function of several key tissues. Included in these tissues are the wool follicles, which have a high demand for glucose (32).
Volatile? Fatty acids
On pastures where the nongluconeogenic volatile fatty acid acetate predominates, sheep are prone to ketonuria and a suboptimal growth of wool (33,34). It has been suggested that if the abundant supply of acetate in these animals could be utilized for glucose production, some of these problems might be overcome. Acetate can serve as a source of glucose in organisms that possess the enzymes necessary to catalyze the reactions of the glyoxylate cycle (35) (Fig. 2) where isocitric acid from the tricarboxylic acid cycle is cleaved to succinate and glyoxylate by the action of the enzyme isocitrate lyase. Succinate is a gluconeogenic substrate, whereas the glyoxylate produced in the reaction is combined with another molecule of acetate to produce malate, thus providing the necessary substrate for continuation of the cycle. This second reaction is catalyzed by the enzyme malate synthase.
In E. coli, the enzyme isocitrate lyase is encoded by the gene aceA, whereas malate synthase is encoded by the aceB gene. These genes
Activity of Isocitrate Lyase and Malate Synthase in Extracts of Zinc-Induced L-Cells Transformed with Fusion Genes Encoding the Enzymes of the Glyoxylate Cycle"
Malate synthase pMTaceAl pMTaceBl pMTaceA2 pMTaceB2
"pMTaceAl and pMTaceBl contain the complete sheep growth-hormone sequence, whereas pMTaceA2 and pMTaceB2 contain only exon 5, as shown in Fig 1 Specific activities are expressed as nmoles of product formed /20 min /mg protein and are corrected for a low level of malate synthase activity in L-cell control extracts No isocitrate lyase activity was detected in untransformed L-cells have both recently been isolated and sequenced (36-38), and their modification for transcription in eukaryotes has been carried out in similar fashion to that described for the cysteine genes, using the sheep MT-Ia promoter sequence and exon 5 of the sheep growth-hormone gene. When transferred to L-cells in culture, all three genes produced RNA transcripts of the predicted sizes (27,39), and these were translated into active enzyme as indicated by the activities of isocitrate lyase and malate synthase in extracts prepared from the transformed cells (Table 3). The genes were then transferred to mice, and their expression examined in liver, kidney, and intestinal tissues, where mRNAs were detected that hybridized with appropriate probes for the bacterial coding sequences and were of the predicted sizes (39). Cell-free extracts from the same tissues showed active isocitrate lyase and malate synthase activities, indicating that the animals have the enzymic potential for the operation of the cycle (39). The detailed physiology of these animals is currently under investigation.
Was this article helpful?