Al-Saleh S, Gore M, Akhtar M
Department of Biochemistry, University of Southampton, U.K.
Biochem J. 1987 Aug 15;246(1):131-7. doi: 10.1042/bj2460131.
Carboxymethylation using 14C- or 3H-labelled iodoacetic acid has been used to identify the cysteine residues in bovine rhodopsin involved in the formation of the two intramolecular disulphide bridges. Iodo[2-14C]acetic acid was used to modify 5.8-5.9 residues of cysteine under non-reducing conditions. After dialysis and reduction of disulphide bridges by 2-mercaptoethanol, iodo[2-3H]acetic acid was employed to covalently modify 3.3-3.6 residues of cysteine. Peptide purification and sequencing has unambiguously shown that cysteine residues 322 and 323 are only carboxymethylated after reduction of disulphide bridges. Indirect evidence presented, now coupled with the earlier finding [Findlay & Pappin (1986) Biochem. J. 238, 625-642] suggests that the other disulphide bridge is formed between cysteine residues 110 and 187. A comparison is made of all the sequences of mammalian rhodopsins and colour pigments and attention is drawn to the fact that whereas Cys-322 and Cys-323 are conserved only in three rhodopsins (bovine, ovine and human), the residues corresponding to Cys-110 and Cys-187 are found in all the visual proteins (from rods as well as human cones).
使用14C或3H标记的碘乙酸进行羧甲基化，已被用于鉴定牛视紫红质中参与形成两个分子内二硫键的半胱氨酸残基。在非还原条件下，碘[2-14C]乙酸用于修饰5.8 - 5.9个半胱氨酸残基。在通过2-巯基乙醇透析并还原二硫键后，碘[2-3H]乙酸用于共价修饰3.3 - 3.6个半胱氨酸残基。肽的纯化和测序明确表明，只有在二硫键还原后，半胱氨酸残基322和323才会被羧甲基化。所提供的间接证据，现在与早期的发现[芬德利和帕平（1986年）《生物化学杂志》238卷，625 - 642页]相结合，表明另一个二硫键是在半胱氨酸残基110和187之间形成的。对所有哺乳动物视紫红质和色色素的序列进行了比较，并注意到这样一个事实：虽然半胱氨酸-322和半胱氨酸-323仅在三种视紫红质（牛、羊和人）中保守，但与半胱氨酸-110和半胱氨酸-187对应的残基存在于所有视觉蛋白中（来自视杆细胞以及人类视锥细胞）。
