Protein succination is a post-translational modification formed by a reaction between the Krebs cycle intermediate fumarate with protein cysteines to form S-(2-succino)cysteine (2SC). Cysteine predominantly exists in the thiolate form and can act as a reactive nucleophile. Cysteine succination can occur non-enzymatically and is formed by a Michael Addition reaction between fumarate and the free thiol groups of protein cysteines at physiological pH. The thioether bond of 2SC is considered to be stable to acid hydrolysis and irreversible. Fumarate is a weak electrophile and its modification of thiols is highly pH dependent. As a consequence, succination can be selective towards functional, low pKa cysteine residues in proteins, such as catalytic cysteine residues in enzymes.
Figure 1: Diagram showing the Michael Addition reaction to form S-(2-succinocysteine)
Succination at critical cysteine residues can result in the inactivation of enzymatic activity or protein function in many biological processes. For example, the succination of key components of the iron-sulfur cluster biogenesis family of proteins, Iscu and Nfu1, lead to defects in iron-sulfur biosynthesis required for respiratory chain complexes. Succination of Glutathione has been shown to increase oxidative stress and cellular senescence. The loss of Fumarate Hydratase (FH), the enzyme that catalyzes the reversible hydration/dehydration of fumarate to L-malate, contributes to the accumulation of fumarate and therefore succination. FH deficiency leads to the inactivation of the E3 ubiquitin ligase Keap1 (Kelch-like ECH-associated protein 1) by succination, which promotes the stabilization of Nuclear Factor 2 (NRF2) and activation of the antioxidant pathway. Keap1 also plays a key role in controlling tumorigenesis.
2SC is considered as a biomarker for mitochondrial stress in obesity, insulin resistance and diabetes. The succination of adiponectin is increased in adipocytes and adipose tissue of type 2 diabetic mice. Adiponectin succination blocks the formation of oligomeric species and secreted forms of adiponectin, which contributes to reduced levels of plasma adiponectin in diabetes. Succination causes irreversible inactivation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) resulting in the loss of activity in muscle of diabetic rats. The elucidation of the succinated proteome will provide an insight into the role of succination in regulatory biology and determine its effect on cellular dysfunction.
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Figure 2: Western blot analysis of mouse epithelial kidney (MEK) fumarate hydratase Fh1 fl/fl wildtype and Fh1 -/- knockout cell lysates preparation without and with OVA-2SC antigen. Lane 1: MEK Fh1 fl/fl (100μg) Lane 2: MEK Fh1 -/- (100µg)
For more information see:
Frizzell, N. et al (2013). The succinated proteome. Mass Spectrometry Reviews, 33(2), pp.98-109. PMID: PMC4038156
Frizzell, N. et al. (2007). Succination of Protein Thiols during Adipocyte Maturation: A BIOMARKER OF MITOCHONDRIAL STRESS. Journal of Biological Chemistry, 282(47), pp.34219-34228. PMID: 17726021