Quiroz-Valenzuela Soledad, Sukuru Sai Chetan K, Hausinger Robert P, Kuhn Leslie A, Heller William T
Department of Biochemistry & Molecular Biology, Michigan State University, East Lansing, MI 48824, USA.
Arch Biochem Biophys. 2008 Dec 1;480(1):51-7. doi: 10.1016/j.abb.2008.09.004. Epub 2008 Sep 18.
Conformational changes of Klebsiella aerogenes urease apoprotein (UreABC)(3) induced upon binding of the UreD and UreF accessory proteins were examined by a combination of flexibility analysis, mutagenesis, and small-angle X-ray scattering (SAXS). ProFlex analysis of urease provided evidence that the major domain of UreB can move in a hinge-like motion to account for prior chemical cross-linking results. Rigidification of the UreB hinge region, accomplished through a G11P mutation, reduced the extent of urease activation, in part by decreasing the nickel content of the mutant enzyme, and by sequestering a portion of the urease apoprotein in a novel activation complex that includes all of the accessory proteins. SAXS analyses of urease, (UreABC-UreD)(3), and (UreABC-UreDF)(3) confirm that UreD and UreF bind near UreB at the periphery of the (UreAC)(3) structure. This study supports an activation model in which a domain-shifted UreB conformation in (UreABC-UreDF)(3) allows CO(2) and nickel ions to gain access to the nascent active site.
通过灵活性分析、诱变和小角X射线散射（SAXS）相结合的方法，研究了产气克雷伯菌脲酶脱辅基蛋白（UreABC）(3)在结合UreD和UreF辅助蛋白时诱导产生的构象变化。脲酶的ProFlex分析提供了证据，表明UreB的主要结构域可以以类似铰链的运动方式移动，以解释先前的化学交联结果。通过G11P突变实现的UreB铰链区的刚性化，部分通过降低突变酶的镍含量，并通过将一部分脲酶脱辅基蛋白隔离在一个包括所有辅助蛋白的新型活化复合物中，从而降低了脲酶活化的程度。对脲酶、(UreABC-UreD)(3)和(UreABC-UreDF)(3)的SAXS分析证实，UreD和UreF在(UreAC)(3)结构的外围靠近UreB处结合。本研究支持一种活化模型，其中(UreABC-UreDF)(3)中结构域移位的UreB构象允许CO(2)和镍离子进入新生的活性位点。
