Ozolin' O N, Uteshev T A, Kim Iu A, Deev A A, Kamzolova S G
Mol Biol (Mosk). 1990 Jul-Aug;24(4):1057-66.
The method for specific modification of Escherichia coli RNA polymerase by a monomercuric derivative of fluorescein--fluoresceinmonomercuracetate (FMMA)--a specific reagent for SH-groups of proteins is suggested. It is shown, that in conditions of equimolar FMMA/enzyme ratio the fluorescent label interacts preferantially with a single sulfhydryl group in alpha-subunit of RNA polymerase. The mercaptide bonding formation is followed by significant alterations of all spectral parameters of FMMA, but has no effect on the kinetic parameters (KB and k2) of RNA synthesis initiation nor does it lead to inhibition of the total RNA synthesis. The modification presented may be used in structural and topological investigations of RNA polymerase functioning.
提出了一种用荧光素的单汞衍生物——荧光素单巯基乙酸汞(FMMA)对大肠杆菌RNA聚合酶进行特异性修饰的方法,FMMA是一种蛋白质SH基团的特异性试剂。结果表明,在FMMA/酶等摩尔比的条件下,荧光标记优先与RNA聚合酶α亚基中的一个巯基相互作用。硫醇盐键的形成伴随着FMMA所有光谱参数的显著变化,但对RNA合成起始的动力学参数(KB和k2)没有影响,也不会导致总RNA合成的抑制。所提出的修饰可用于RNA聚合酶功能的结构和拓扑研究。