4‑Hydroxy-7-oxo-5-heptenoic
Acid (HOHA) Lactone
is a Biologically Active Precursor for the Generation of 2‑(ω-Carboxyethyl)pyrrole
(CEP) Derivatives of Proteins and Ethanolamine Phospholipids
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Abstract
2-(ω-Carboxyethyl)pyrrole
(CEP) derivatives of proteins were
previously shown to have significant pathological and physiological
relevance to age-related macular degeneration, cancer and wound healing.
Previously, we showed that CEPs are generated in the reaction of ε-amino
groups of protein lysyl residues with 1-palmityl-2-(4-hydroxy-7-oxo-5-heptenoyl)-<i>sn</i>-glycero-3-phosphatidylcholine (HOHA-PC), a lipid oxidation
product uniquely generated by oxidative truncation of docosahexanenate-containing
phosphatidylcholine. More recently, we found that HOHA-PC rapidly
releases HOHA-lactone and 2-lyso-PC (<i>t</i><sub>1/2</sub> = 30 min at 37 °C) by nonenzymatic transesterification/deacylation.
Now we report that HOHA-lactone reacts with Ac-Gly-Lys-OMe or human
serum albumin to form CEP derivatives in vitro. Incubation of human
red blood cell ghosts with HOHA-lactone generates CEP derivatives
of membrane proteins and ethanolamine phospholipids. Quantitative
analysis of the products generated in the reaction HOHA-PC with Ac-Gly-Lys-OMe
showed that HOHA-PC mainly forms CEP-dipeptide that is not esterified
to 2-lysophosphatidycholine. Thus, the HOHA-lactone pathway predominates
over the direct reaction of HOHA-PC to produce the CEP-PC-dipeptide
derivative. Myleoperoxidase/H<sub>2</sub>O<sub>2</sub>/NO<sub>2</sub><sup>–</sup> promoted in vitro oxidation of either 1-palmityl-2-docosahexaneoyl-<i>sn</i>-glycero-3-phosphatidylcholine (DHA-PC) or docosahexaenoic
acid (DHA) generates HOHA-lactone in yields of 0.45% and 0.78%, respectively.
Lipid oxidation in human red blood cell ghosts also releases HOHA-lactone.
Oxidative injury of ARPE-19 human retinal pigmented epithelial cells
by exposure to H<sub>2</sub>O<sub>2</sub> generated CEP derivatives.
Treatment of ARPE-19 cells with HOHA-lactone generated CEP-modified
proteins. Low (submicromolar), but not high, concentrations of HOHA-lactone
promote increased vascular endothelial growth factor (VEGF) secretion
by ARPE-19 cells. Therefore, HOHA-lactone not only serves as an intermediate
for the generation of CEPs but also is a biologically active oxidative
truncation product from docosahexaenoate lipids