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Dhinakar S. Kompala
Research Interests

Recombinant mammalian cell cultures:
We have developed a novel inducible expression system for overproducing glycoproteins in recombinant mammalian cells, such as the CHO and BHK cells. This system utilizes the co-expression of a specific transcription factor, glucocorticoid receptor, for the inducible expression of a reporter glycoprotein from the mouse mammary tumor virus (MMTV) promoter. We have shown that this expression system produces an order of magnitude higher expression of the reporter glycoprotein, secreted alkaline phosphatase, after induction by a glucocorticoid analog, compared to the typical expression from constitutive promoters, such as the CMV and SV40 promoters. In our current efforts to characterize and further optimize the genetic regulatory network in this runaway, positive feedback expression system, we are utilizing modern molecular biotechnology tools, such as subtractive suppressive hybridization, representational difference analysis 2D gel proteomics, and MALDI-TOF mass spectrometry.

Metabolic engineering of Zymomonas mobilis to ferment pentose sugars:
In this collaborative project with the Biotechnology Center for Fuels and Chemicals at the National Renewable Energy Laboratory in Golden, Colorado, we are investigating the effects of gene dosage and controlled gene expression in maximizing the metabolic flux along the newly engineering pentose utilization pathway. We are characterizing the dynamic enzymatic activity levels for each newly incorporated enzymes and a few key native glycolytic pathway enzymes. Strong variations in the key enzymatic activity levels are being further investigated through 2D gel electrophoresis techniques. Intracellular key metabolite concentrations are being monitored through NMR spectroscopy. These dynamic measurements will be used to develop dynamic models of the metabolic fluxes through the different substrate utilization pathways and later to maximize the ethanol production rate in various bioreactor operating strategies.

Metabolic oscillations in the yeast Saccharomyces cerevisiae:
In this new research project, we address the growing controversy on the mechanistic causes for the sustained oscillations over some range of parameters in the chemostat cultures of yeast cells. While the experimentally observed partial cell cycle synchrony has been hypothesized as the causative mechanism by some others, our dynamic simulations of the 3 major catabolic pathways in yeast suggest that these oscillations are caused by the regulatory processes at the gene expression level. To investigate the this mechanistic cause, we are currently embarking on DNA microarray analysis of gene expression of yeast cells from different chemostat oscillation regimes and correlating the oscillations in gene expression with the known metabolic functions.