Electrospray Ionisation Studies of the Interaction of N,N′-Dicyclohexylcarbodiimide with Ceroid Lipofuscinosis Protein (Subunit C)

N,N′-dicyclohexylcarbodiimide (DCCD) interacts with isolated ceroid lipofuscinosis protein (CLP), a model for subunit c. The products are (a) CLP–DCCD adduct (+206 Da), presumably by addition at Glu-58; (b) acetyl–CLP–DCCD (+42 Da + 206 Da); (c) acetyl–CLP (+42 Da). Under specific conditions, additional DCCD adducts are formed; CLP–DCCD–DCCD (+206 Da + 206 Da) and acetyl–CLP–DCCD–DCCD (+42 Da + 206 Da + 206 Da). Acetylation utilises the acetate present in CLP preparations as a buffer and the site is shown to be located within amino acids 1–9 (probably Lys-7 or N-terminal aspartate). Acetyl–CLP is shown to be the primary product but, in addition, acylation by other fatty acids (myristate, palmitate, oleate and stearate) can be demonstrated in low acetate concentrations. Oligomycin is shown to modify some interactions of CLP with DCCD but venturicidin has little or no effect.