Linked Life Data

19645454

Source:http://linkedlifedata.com/resource/pubmed/id/19645454

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
36
pubmed:dateCreated
2009-9-8
pubmed:abstractText
Allosteric regulation of human lipoxygenase (hLO) activity has recently been implicated in the cellular biology of prostate cancer. In the current work, we present isotope effect, pH, and substrate inhibitor data of epithelial 15-hLO-2, which probe the allosteric effects on its mechanistic behavior. The Dk(cat)/KM for 15-hLO-2, with AA and LA as substrate, is large indicating hydrogen atom abstraction is the principle rate-determining step, involving a tunneling mechanism for both substrates. For AA, there are multiple rate determining steps (RDS) at both high and low temperatures, with both diffusion and hydrogen bonding rearrangements contributing at high temperature, but only hydrogen bonding rearrangements contributing at low temperature. The observed kinetic dependency on the hydrogen bonding rearrangement is eliminated upon addition of the allosteric effector, 13-(S)-hydroxyoctadecadienoic acid (13-HODE), and no allosteric effects were seen on diffusion or hydrogen atom abstraction. The (k(cat)/KM)AA/(k(cat)/KM)LA ratio was observed to have a pH dependence, which was fit with a titration curve (pKa = 7.7), suggesting the protonation of a histidine residue, which could hydrogen bond with the carboxylate of 13-HODE. Assuming this interaction, 13-HODE was docked to the solvent exposed histidines of a 15-hLO-2 homology model and found to bind well with H627, suggesting a potential location for the allosteric site. Utilizing d31-LA as an inhibitor, it was demonstrated that the binding of d31-LA to the allosteric site changes the conformation of 15-hLO-2 such that the affinity for substrate increases. This result suggests that allosteric binding locks the enzyme into a catalytically competent state, which facilitates binding of LA and decreases the (k(cat)/KM)AA/(k(cat)/KM)LA ratio. Finally, the magnitude of the 13-HODE KD for 15-hLO-2 is over 200-fold lower than that of 13-HODE for 15-hLO-1, changing the substrate specificity of 15-hLO-2 to 1.9. This would alter the LO product distribution and increase the production of the pro-tumorigenic, 13-HODE, possibly representing a pro-tumorigenic feedback loop for 13-HODE and 15-hLO-2.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
1520-4995
pubmed:author
pubmed:issnType
Electronic
pubmed:day
15
pubmed:volume
48
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
8721-30
pubmed:dateRevised
2011-9-26
pubmed:meshHeading
pubmed-meshheading:19645454-Allosteric Site, pubmed-meshheading:19645454-Arachidonate 15-Lipoxygenase, pubmed-meshheading:19645454-Arachidonic Acid, pubmed-meshheading:19645454-Binding, Competitive, pubmed-meshheading:19645454-Deuterium Exchange Measurement, pubmed-meshheading:19645454-Epithelial Cells, pubmed-meshheading:19645454-Histidine, pubmed-meshheading:19645454-Humans, pubmed-meshheading:19645454-Isoenzymes, pubmed-meshheading:19645454-Kinetics, pubmed-meshheading:19645454-Lipoxygenase Inhibitors, pubmed-meshheading:19645454-Male, pubmed-meshheading:19645454-Models, Molecular, pubmed-meshheading:19645454-Prostate, pubmed-meshheading:19645454-Protein Binding, pubmed-meshheading:19645454-Solvents, pubmed-meshheading:19645454-Structural Homology, Protein, pubmed-meshheading:19645454-Substrate Specificity, pubmed-meshheading:19645454-Tandem Mass Spectrometry
pubmed:year
2009
pubmed:articleTitle
Kinetic and structural investigations of the allosteric site in human epithelial 15-lipoxygenase-2.
pubmed:affiliation
Chemistry and Biochemistry Department, University of California, Santa Cruz, California 95064, USA.
pubmed:publicationType
Journal Article, Comparative Study, Research Support, Non-U.S. Gov't, Research Support, N.I.H., Extramural