Journal
JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY
Volume 21, Issue 5-6, Pages 605-618Publisher
SPRINGER
DOI: 10.1007/s00775-016-1373-8
Keywords
Nonheme diiron enzymes; Oxygen activation; Peroxo intermediates; Deoxyhypusine hydroxylase; Cell death
Funding
- National Institutes of Health [GM-38767, 5F32GM106612-02]
- Intramural Research Program of the National Institute of Dental and Craniofacial Research
- Vietnam Education Foundation
- US Department of Energy (DOE), Office of Science, Office of Basic Energy Sciences [DE-AC02-76SF00515]
- DOE Office of Biological and Environmental Research
- National Institutes of Health, National Institute of General Medical Sciences [P41GM103393]
- DOE Office of Science by Brookhaven National Laboratory [DE-AC02-98CH10886]
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Human deoxyhypusine hydroxylase (hDOHH) is an enzyme that is involved in the critical post-translational modification of the eukaryotic translation initiation factor 5A (eIF5A). Following the conversion of a lysine residue on eIF5A to deoxyhypusine (Dhp) by deoxyhypusine synthase, hDOHH hydroxylates Dhp to yield the unusual amino acid residue hypusine (Hpu), a modification that is essential for eIF5A to promote peptide synthesis at the ribosome, among other functions. Purification of hDOHH overexpressed in E. coli affords enzyme that is blue in color, a feature that has been associated with the presence of a peroxo-bridged diiron(III) active site. To gain further insight into the nature of the diiron site and how it may change as hDOHH goes through the catalytic cycle, we have conducted X-ray absorption spectroscopic studies of hDOHH on five samples that represent different species along its reaction pathway. Structural analysis of each species has been carried out, starting with the reduced diferrous state, proceeding through its O-2 adduct, and ending with a diferric decay product. Our results show that the Feai-Fe distances found for the five samples fall within a narrow range of 3.4-3.5 , suggesting that hDOHH has a fairly constrained active site. This pattern differs significantly from what has been associated with canonical dioxygen activating nonheme diiron enzymes, such as soluble methane monooxygenase and Class 1A ribonucleotide reductases, for which the Feai-Fe distance can change by as much as 1 during the redox cycle. These results suggest that the O-2 activation mechanism for hDOHH deviates somewhat from that associated with the canonical nonheme diiron enzymes, opening the door to new mechanistic possibilities for this intriguing family of enzymes.
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