Eric L. Hegg Research Interests
1. Elucidating the biosynthesis, transport, and regulation of heme A.
Cytochrome c oxidase (CcO) is the terminal oxidase in all plants, animals, aerobic yeasts, and many bacteria. Arguably the most important enzyme in aerobic metabolism, CcO catalyzes the transfer of electrons 
from cytochrome c to O2 concomitant with the pumping of protons across the inner membrane of either mitochondria or bacteria. This proton gradient is ultimately utilized to generate approximately 50% of the ATP formed during aerobic metabolism. The site of O2 reduction is an unusual heterobimetallic center consisting of a copper ion and a modified heme known as heme A. The biosynthesis of heme A and its insertion into CcO is a complex process that is only just beginning to be elucidated at the molecular level. Heme A is derived from heme B via two enzymatic reactions. The first reaction, catalyzed by heme O synthase (HOS), results in the conversion of the vinyl group on pyrrole ring A into a 17-hydroxyethylfarnesyl moiety. In the second transformation, heme A synthase (HAS) utilizes a heme B cofactor to catalyze the oxidation of the methyl group on pyrrole ring D into an aldehyde. The second reaction is especially intriguing because this is an unusual oxidation reaction in biological systems, and the enzyme that catalyzes this reaction is itself a heme-containing enzyme.
The focus of our research is to elucidate the biosynthesis of heme A and its method of transport, including the formation of associated multi-component complexes. Because free heme is toxic to cells, the regulation and transport of hemes must be rigorously controlled, and our lab is providing important insight into how this occurs. To address these and related questions, we work with a number of different prokaryotic and eukaryotic organisms including E. coli, B. subtilis, R. sphaeroides, and S. cerevisiae. In doing so, we not only elucidate the biosynthetic mechanism of a key metabolic cofactor and enhance our understanding of how nature uses O2 to oxidize the very stable C-H bond, we also obtain fundamental information concerning heme transport and homeostasis.
2. Identifying and characterizing novel hydrogenase enzymes from diverse organisms including both halophiles and thermophiles.
Hydrogen has enormous potential to serve as a renewable and non-polluting fuel. Not surprisingly, Nature has already discovered the potential in H2 and has developed a class of enzymes (hydrogenases) which are among the most efficient H2-generating catalysts known. Hydrogenases are found throughout nature in many taxonomically diverse microorganisms, illustrating the critical role of H2 metabolism in the world's microbial environments.
Several microbes hold significant promise for the commercial production of H2. Photosynthetic microalgae are unique in that H2 production can be coupled directly to water oxidation through photosystem II and the photosynthetic electron transport chain, providing the means to generate H2 directly from sunlight and water.
Currently there are a number of major hurdles that must be overcome before the bio-production of H2 using photosynthetic microalgae is commercially viable. Perhaps is the biggest hurdle is that hydrogenase enzymes are inactivated in the presence of O2 - an unfortunate and ironic impediment to our quest of coupling hydrogen production to photosynthesis. As part of a multi-disciplinary team (http://www.princeton.edu/~biosolar/), our goal is to utilize a combination of bioprospecting and targeted searching to identify and to characterize novel and unusually active/stable hydrogenases for use in biosolar hydrogen production.
3. Assessing metabolic activity by monitoring the isotope ratio of intracellular water.
The transport of water into and out of cells is a seemingly simple process in which water diffuses either through pores or directly across the membrane. Although polar molecules are generally unable to diffuse across biological membranes, the small size of water allows it to move through defects in the membrane as the lipids diffuse laterally. In addition, water can also be transported through channels called aquaporins at essentially diffusion-controlled rates. The rate at which these two processes can theoretically occur has led to the generally accepted assumption that intracellular water is isotopically indistinguishable from extracellular water.
Using stable isotope ratio mass spectrometry we directly tested this assumption, and our data indicate that this commonly held assumption is NOT correct. In fact, during periods of active growth, a large percentage of the intracellular water can be isotopically distinct from extracellular water due to the water that is generated during metabolism. Significantly, the amount of intracellular water that is the product of metabolism is a function of metabolic rate. Therefore, we can assess the total metabolic rate of cells by measuring the isotope ratio of intracellular water. In addition, the isotope ratio of metabolites (e.g., fatty acids) that incorporate either oxygen or hydrogen atoms from water during biosynthesis can also be used as a probe of metabolic rate and therefore as a probe of metabolic status.
Our ability to assess the metabolic activity of cells by measuring the isotope ratio of intracellular water or other cellular components has a number of potentially interesting applications. These applications include:
- Cancer diagnostic - determining how fast a cell is metabolizing.
- Obesity research - elucidating what conditions stimulate (or inhibit) metabolism on a cellular level.
- Hydrogen/energy research - elucidating the flow of protons through hydrogen-evolving organisms.
- Environmental and health research - ascertaining the heath of colonies by studying the metabolic rate of "mats" or other biofilms.