Non-oxidative modification of lens crystallins by kynurenine: A novel post-translational protein modification with possible relevance to ageing and cataract

Brett Garner*, Denis C. Shaw, Robyn A. Lindner, John A. Carver, Roger J.W. Truscott

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    72 Citations (Scopus)

    Abstract

    In humans, the crystallin proteins of the ocular lens become yellow- coloured and fluorescent with ageing. With the development of senile nuclear cataract, the crystallins become brown and additional fluorophores are formed. The mechanism underlying crystallin colouration is not known but may involve interaction with kynurenine-derived UV filter compounds. We have recently identified a sulphur-linked glutathionyl-3-hydroxykynurenine glucoside adduct in the lens and speculated that kynurenine may also form adducts with GSH and possibly with nucleophilic amino acids of the crystallins (e.g. Cys). Here we show that kynurenine modifies calf lens crystallins non-oxidatively to yield coloured (365 nm absorbing), fluorescent (Ex 380 nm/Em 450-490 nm) protein adducts. Carboxymethylation and succinylation of crystallins inhibited kynurenine-mediated modification by approx. 90%, suggesting that Cys, Lys and possibly His residues may be involved. This was confirmed by showing that kynurenine formed adducts with GSH as well as with poly-His and poly-Lys. NMR studies revealed that the novel poly-Lys-kynurenine covalent linkage was via the ε-amino group of the Lys side chain and the βC of the kynurenine side chain. Analysis of tryptic peptides of kynurenine-modified crystallins revealed that all of the coloured peptides contained either His, Cys or an internal Lys residue. We propose a novel mechanism of kynurenine-mediated crystallin modification which does not require UV light or oxidative conditions as catalysts. Rather, we suggest that the side chain of kynurenine-derived lens UV filters becomes deaminated to yield an α,β-unsaturated carbonyl which is highly susceptible to attack by nucleophilic amino acid residues of the crystallins. The inability of the lens fibre cells to metabolise their constituent proteins results in the accumulation of coloured/fluorescent crystallins with age. (C) 2000 Elsevier Science B.V.

    Original languageEnglish
    Pages (from-to)265-278
    Number of pages14
    JournalBiochimica et Biophysica Acta - Protein Structure and Molecular Enzymology
    Volume1476
    Issue number2
    DOIs
    Publication statusPublished - 9 Feb 2000

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