Krueger Sharon K, Henderson Marilyn C, Siddens Lisbeth K, VanDyke Jonathan E, Benninghoff Abby D, Karplus P Andrew, Furnes Bjarte, Schlenk Daniel, Williams David E
Linus Pauling Institute, Oregon State University, Corvallis, OR 97331, USA.
Drug Metab Dispos. 2009 Aug;37(8):1785-91. doi: 10.1124/dmd.109.027201. Epub 2009 May 6.
Catalytically active human flavin-containing monooxygenase isoform 2 (FMO2.1) is encoded by an allele detected only in individuals of African or Hispanic origin. Genotyping and haplotyping studies indicate that S195L and N413K occasionally occur secondary to the functional FMO21 allele encoding reference protein Gln472. Sulfoxygenation under a range of conditions reveals the role these alterations may play in individuals expressing active FMO2 and provides insight into FMO structure. Expressed S195L lost rather than gained activity as pH was increased or when cholate was present. The activity of S195L was mostly eliminated after heating at 45 degrees C for 5 min in the absence of NADPH, but activity was preserved if NADPH was present. By contrast, Gln472 was less sensitive to heat, a response not affected by NADPH. A major consequence of the S195L mutation was a mean 12-fold increase in K(m) for NADPH compared with Gln472. Modeling an S213L substitution, the equivalent site, in the structural model of FMO from the Methylophaga bacterium leads to disruption of interactions with NADP(+). N413K had the same pattern of activity as Gln472 in response to pH, cholate, and magnesium, but product formation was always elevated by comparison. N413K also lost more activity when heated than Gln472; however, NADPH attenuated this loss. The major effects of N413K were increases in velocity and k(cat) compared with Gln472. Although these allelic variants are expected to occur infrequently as mutations to the FMO21 allele, they contribute to our overall understanding of mammalian FMO structure and function.
具有催化活性的人含黄素单加氧酶同工型2（FMO2.1）由仅在非洲裔或西班牙裔个体中检测到的一个等位基因编码。基因分型和单倍型研究表明，S195L和N413K偶尔会继发于编码参考蛋白Gln472的功能性FMO21等位基因。在一系列条件下的硫氧化反应揭示了这些改变可能在表达活性FMO2的个体中所起的作用，并为FMO结构提供了深入了解。随着pH值升高或存在胆酸盐时，表达的S195L失去而非获得活性。在不存在NADPH的情况下，S195L在45℃加热5分钟后活性大多被消除，但如果存在NADPH则活性得以保留。相比之下，Gln472对热不太敏感，这种反应不受NADPH影响。S195L突变的一个主要后果是与Gln472相比，NADPH的K(m)平均增加了12倍。在来自嗜甲基菌的FMO结构模型中模拟等效位点的S213L取代会导致与NADP(+)的相互作用中断。N413K在对pH、胆酸盐和镁的反应中具有与Gln472相同的活性模式，但相比之下产物形成总是升高。与Gln472相比，N413K在加热时也失去了更多活性；然而，NADPH减弱了这种损失。与Gln472相比，N413K的主要影响是速度和k(cat)增加。尽管预计这些等位基因变体作为FMO21等位基因的突变很少发生，但它们有助于我们对哺乳动物FMO结构和功能的整体理解。
