Characterization of Aldolase and Transaldolase Activities in Acidobacterium capsulatum

Abstract

Acidobacterium capsulatum, a mildly acidophilic, chemotrophic, aerobe with the ability to grow on glucose, is found in diverse soil and aquatic conditions worldwide, suggesting a significant role in carbon and nutrient cycling in the environment. A. capsulatum has successfully been cultured and its genome has been completed, allowing for genome-directed studies of the organism's glucose metabolizing capabilities. Based on the genomic information, this organism was thought to lack aldolase, but contained two putative transaldolase genes (TA), one of which contains and EF-hand domain (TAEF). However, recent results show that there are both aldolase and transaldolase activities in cell-free lysate. We are continuing to characterize the activities of aldolase and transaldolase in cell-free lysate to better understand partitioning of carbon from glucose to energy production and biomass formation. Characterization will include pH and EDTA dependent studies to determine aldolase and transaldolase activity dependence on pH and calcium. Furthermore the two putative transaldolase genes have been amplified, cloned into E. coli and transformed into a pET151 directional TOPO expression vector. Experiments to determine whether the transaldolase genes can be expressed are in progress. Sequencing of the TAEF gene showed proper insertion into the expression vector. Current work is focusing on purification and expression of the TA and TAEF proteins. The evidence from these studies will allow for a better understanding of the role of transaldolase and aldolase as mediators of carbon partitioning between energy production and biomass formation in A. capsulatum. Future work will focus on identifying an aldolase gene and characterizing key regulatory points of glucose metabolism.