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theses

During the past five decades, mercury has gained increased interest due to its toxicity to human and environmental health. Therefore, mercury detoxification, whereby the mercuric reductase (MR), a homodimer of MerA (Figure 3 10), converts Hg2+ to Hg0, is an important activity. merA, the gene encoding MerA, has been found in diverse Archaea and Bacteria [1], but it is not well known in the Bacteroidetes, a large phylum in the bacterial domain that is widely distributed in many environments. The goal of this study was to identify protein structural characteristics that relate to MerA temperature optimum for activity in two species of the phylum Bacteroidetes: one a thermophile, Rhodothermus marinus, and the other a psychrophile, Flavobacterium. sp. SOK62. The standard MerA assay [2] was optimized by adjusting pH, selecting the reducing substrates (NADH/NADPH) and the type and concentration of thiol agent. Using the optimized assay, I found that the optimum temperature for MerA of R. marinus was at 65-70˚C (activity range from 30 to 90 ºC) and for the psychrophilic MerA (strain SOK62) was at 50-55 ˚C (range from 10 to 90 ºC). Homology modeling (Figure 3 7) of the psychrophilic and thermophilic MerA (homology to a proteobacterial MerA from Pseudomonas aeruginosa PAT) showed that the psychrophile’s MerA (SOK62) has more α-helix and less β-sheet secondary structure than the thermophile’s MerA (R. marinus), which is shown in Table 3 5. MerA of SOK62 has more polar residues and less hydrophobic residues, suggesting adaptation to activity at lower temperatures [3]than MerA of R. marinus. In contrast, the psychrophile’s MerA has a larger number of aromatic residues than the R. marinus enzyme, contradicting the expectation of a lower number of bulky residues in a psychrophilic protein. These experiments test the hypothesis that because MerA originated among thermophiles in geothermal environments [4], the MerA from a psychrophilic bacterium has a thermophilic enzyme activity optimum and structural adaptations facilitating activity at low temperatures. This study contributes to our understanding of the natural history of microbial mercury detoxification.

ETD_5858

M.S.

Includes bibliographical references

by Bahram Manavi

doi:10.7282/T3QZ28NV

ETD graduate

The author owns the copyright to this work.