A one-carbon modification of protein lysine associated with elevated oxidative stress in human substantia nigra.
We describe for the first time a naturally occurring lysine modification that is converted to methyllysine by reduction with sodium borohydride. This modification is approximately 1.7 times as abundant in soluble proteins from human substantia nigra pars compacta as in proteins from other brain regions, possibly as a result of elevated oxidative stress in the nigra. Proteins from cultured PC12 cells exposed to oxidative stress conditions also contain elevated levels of this lysine modification. The abundance of the naturally occurring modification is roughly 0.08 nmoles/mg protein in either unstressed brain or PC12 cells. Modification levels remain stable in isolated proteins incubated for 2 h at 37 degrees C in pH 7 buffer. We propose that the endogenous modification is the lysine Schiff base, epsilon-N-methylenelysine, and that lysine modifications may result from a reaction with formaldehyde in vivo. Rat brain contains approximately 60 nmoles/g wet weight of formaldehyde, which probably includes both free and reversibly bound forms. Adding approximately 35 microm HCHO to PC12 cell growth medium introduces methylenelysine modifications in cell proteins and impairs cell viability. The existence of this post-translational modification suggests new mechanisms of oxidative stress that may contribute to tissue degeneration, including loss of nigral dopamine neurons during normal aging and in Parkinson's disease.
Floor
Erik
E
Department of Molecular Biosciences and Mass Spectrometry Laboratory, University of Kansas, Lawrence, 66045, USA. floor@ku.edu
Maples
Anne M
AM
Rankin
Carolyn A
CA
Yaganti
Vamsee M
VM
Shank
Sylvan S
SS
Nichols
Grant S
GS
O'Laughlin
Michael
M
Galeva
Nadezhda A
NA
Williams
Todd D
TD
D000368:Aged; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D001894:Borohydrides / Q000493:pharmacokinetics; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D002244:Carbon / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005557:Formaldehyde / Q000378:metabolism / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000494:pharmacology; D006868:Hydrolysis; D008239:Lysine / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D016716:PC12 Cells; D011180:Postmortem Changes; D011499:Protein Processing, Post-Translational / Q000187:drug effects / Q000502:physiology; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D013378:Substantia Nigra / Q000503:physiopathology*; D013997:Time Factors
D000596:Amino Acids; D001894:Borohydrides; D004791:Enzyme Inhibitors; D011506:Proteins; D005557:Formaldehyde; D019328:Buthionine Sulfoximine; D002244:Carbon; D006861:Hydrogen Peroxide; D005978:Glutathione; D008239:Lysine