Lepesheva G I, Hargrove T Y, Ott R D, Nes W D, Waterman M R
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
Biochem Soc Trans. 2006 Dec;34(Pt 6):1161-4. doi: 10.1042/BST0341161.
Sterol 14alpha-demethylases (CYP51) are metabolic cytochromes P450, found in each biological kingdom. They catalyse a single three-step reaction included in all sterol biosynthetic pathways. Plant CYP51s have strict preference towards their physiological substrate O (obtusifoliol), which is C-4-monomethylated. Natural substrates of animal/fungal CYP51 (lanosterol, 24,25-dihydrolanosterol or 24-methylenelanosterol) are C-4-dimethylated. CYP51 from the pathogenic protozoa TB (Trypanosoma brucei) is the first example of O-specific sterol 14alpha-demethylase in non-photosynthetic organisms. Surprisingly, at 83% amino acid identity to the TB orthologue, CYP51 from TC (Trypanosoma cruzi) clearly prefers C-4-dimethylated sterols. Replacement of animal/fungi-like Ile(105) in the B' helix of TC CYP51 with phenylalanine, the residue found in this position in all plant and other trypanosome CYP51s, dramatically increases the ability of the enzyme to metabolize O, converting it into a more plant-like sterol 14alpha-demethylase. A more than 100-fold increase in binding and turnover is observed for the 24-desmethyl analogue of O [N (norlanosterol)], which is found in vivo in procyclic forms of TB and is a good TB CYP51 substrate in vitro. We believe that (i) N is a non-conventional CYP51 substrate, preferred in TB and perhaps other Trypanosomatidae and (ii) functional similarity of TC CYP51 to animal/fungal orthologues is a result of evolutionary convergence (including F105I mutation), leading to different pathways for sterol production in TC versus TB.
甾醇14α-去甲基酶（CYP51）是一种代谢性细胞色素P450，存在于每个生物界。它们催化所有甾醇生物合成途径中包含的一个单一的三步反应。植物CYP51对其生理底物O（ obtusifoliol）有严格的偏好，O是C-4-单甲基化的。动物/真菌CYP51的天然底物（羊毛甾醇、24,25-二氢羊毛甾醇或24-亚甲基羊毛甾醇）是C-4-二甲基化的。致病原生动物布氏锥虫（Trypanosoma brucei）的CYP51是在非光合生物中第一个O特异性甾醇14α-去甲基酶的例子。令人惊讶的是，克氏锥虫（Trypanosoma cruzi）的CYP51与布氏锥虫的直系同源物有83%的氨基酸同一性，但它明显偏好C-4-二甲基化的甾醇。将克氏锥虫CYP51的B'螺旋中类似动物/真菌的异亮氨酸（Ile）（105位）替换为苯丙氨酸（在所有植物和其他锥虫CYP51的该位置发现的残基），会显著提高该酶代谢O的能力，将其转化为更类似植物的甾醇14α-去甲基酶。对于O的24-去甲基类似物[N（去甲羊毛甾醇）]，观察到其结合和周转增加了100多倍，N在布氏锥虫的前循环形式中在体内被发现，并且在体外是布氏锥虫CYP51的良好底物。我们认为：（i）N是一种非常规的CYP51底物，在布氏锥虫以及可能其他锥虫科中更受青睐；（ii）克氏锥虫CYP51与动物/真菌直系同源物的功能相似性是进化趋同的结果（包括F105I突变），导致克氏锥虫与布氏锥虫在甾醇产生途径上有所不同。
