S-(2-Succinyl)cysteine: A novel chemical modification of tissue proteins by a Krebs cycle intermediate

Succination is Increased on Select Proteins in the Brainstem of the NADH dehydrogenase (ubiquinone) Fe-S protein 4 (Ndufs4) Knockout Mouse, a Model of Leigh Syndrome

“…Protein succination refers to the formation of an irreversible thioether linkage between a cysteine residue and the mitochondrial metabolite fumarate (1,2,45). Increased succination in biological samples is restricted to situations where fumarate concentrations are significantly increased, for example, 2SC levels are specifically increased in cancers derived from fumarate hydratase mutations (6,8), and in the metabolically overwhelmed adipocyte in type 2 diabetes (4,5,9).…”

“…The dried sample was resuspended in 1 ml 1% trifluoroacetic acid (TFA) and applied to a C-18 Sep-Pak column (Waters) before elution with 1% TFA/20% methanol. The eluate was dried in vacuo and the residual amino acids were converted to their N,O-trifluoroacetyl methyl ester (TFAME) derivatives for GC-MS/MS analysis as previously described (1,2,12). Quantification was performed by isotope-dilution mass spectrometry based on standard curves.…”

“…wrote the paper. 1 The abbreviations used are: 2SC, S-(2-succino)cysteine; 2D, twodimensional; BS, brainstem; CB, cerebellum; C PE , cysteine pyridylethylation; Crus 1, crus 1 ansiform lobule; C 2SC , cysteine succination by fumarate; Ctx, cortex; ER, endoplasmic reticulum; FN, fastigial nucleus; GC, gas chromatography; Gi, gigantocellular reticular nucleus; GSH, glutathione; HLRCC, hereditary leiomyomatosis and renal cell carcinoma; HNE, 4-hydroxy-2-nonenal; IO, inferior olive; MD, mitochondrial diseases; M OX , methionine oxidation; Mt, microtubular pellet; OB, olfactory bulb; OXPHOS, oxidative phosphorylation; PVDF, polyvinylidene difluoride; RIPA, radioimmuno precipitation assay; SMt, supernatant after tubulin polymerization; SN, sciatic nerve; Str, striatum; Sup, cleared supernatant; VDAC, voltage-dependent anion channel; VN, vestibular nuclei. In the adipocyte the increase in fumarate and succinated proteins develops as a direct result of mitochondrial stress induced by nutrient excess.…”

“…Given the important role of voltage-dependent anion channels isoforms in the exchange of ADP/ATP between the cytosol and the mitochondria, and the already decreased capacity for ATP synthesis in the Ndufs4 KO mice, we propose that the increased protein succination observed in the BS of these animals would further decrease the already compromised mitochondrial function. These data suggest that fumarate is a novel bio- We previously identified the formation of S-(2-succino)cysteine (2SC) 1 (protein succination) as a result of the irreversible reaction of fumarate with reactive cysteine thiols (1,2). Fumarate concentrations are increased during adipogenesis and adipocyte maturation (2,3), and the excess of glucose and insulin leads to augmented protein succination in the adipose tissue of type 2 diabetic mice (4,5).…”

A computational analysis of S-(2-succino)cysteine sites in proteins

“…The adduction of fumaric acid (FA) to the free sulfhydryl group of certain Cys residues in proteins via a Michael-like reaction leads to the formation of S-(2-succino)cysteine (2SC) sites. Originally detected in plasma proteins [10], including albumin, this post-translational Cys modification type (referred as protein succination) has been described in models of diabetes/obesity, fumarate hydratase (FH)-related diseases, and Leigh syndrome. Increased levels of succinated proteins have been found in murine 3T3-L1 adipocytes cultured in high glucose medium (30 mM, compared with a physiological level of 5 mM), as well as in tissues from streptozotocin-treated rats, db/db (leptin receptor deficient), ob/ob (leptin deficient), and diet-induced obese mice [11][12][13][14].…”