Dias Jorge S, Macedo Anjos L, Ferreira Gloria C, Peterson Francis C, Volkman Brian F, Goodfellow Brian J
Rede de Química e Tecnologia, Centro de Química Fina e Biotecnologia, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica, Portugal.
J Biol Chem. 2006 Oct 20;281(42):31553-61. doi: 10.1074/jbc.M605988200. Epub 2006 Aug 10.
Murine p22HBP, a 22-kDa monomer originally identified as a cytosolic heme-binding protein ubiquitously expressed in various tissues, has 27% sequence identity to murine SOUL, a heme-binding hexamer specifically expressed in the retina. In contrast to murine SOUL, which binds one heme per subunit via coordination of the Fe(III)-heme to a histidine, murine p22HBP binds one heme molecule per subunit with no specific axial ligand coordination of the Fe(III)-heme. Using intrinsic protein fluorescence quenching, the values for the dissociation constants of p22HBP for hemin and protoporphyrin-IX were determined to be in the low nanomolar range. The three-dimensional structure of murine p22HBP, the first for a protein from the SOUL/HBP family, was determined by NMR methods to consist of a 9-stranded distorted beta-barrel flanked by two long alpha-helices. Although homologous domains have been found in three bacterial proteins, two of which are transcription factors, the fold determined for p22HBP corresponds to a novel alpha plus beta fold in a eukaryotic protein. Chemical shift mapping localized the tetrapyrrole binding site to a hydrophobic cleft formed by residues from helix alphaA and an extended loop. In an attempt to assess the structural basis for tetrapyrrole binding in the SOUL/HBP family, models for the p22HBP-protoporphyrin-IX complex and the SOUL protein were generated by manual docking and automated methods.
小鼠p22HBP是一种最初被鉴定为在各种组织中普遍表达的胞质血红素结合蛋白的22 kDa单体，与小鼠SOUL（一种在视网膜中特异性表达的血红素结合六聚体）具有27%的序列同一性。与小鼠SOUL每个亚基通过Fe(III)-血红素与组氨酸的配位结合一个血红素不同，小鼠p22HBP每个亚基结合一个血红素分子，且Fe(III)-血红素没有特定的轴向配体配位。利用蛋白质固有荧光猝灭法，测定了p22HBP对血红素和原卟啉-IX的解离常数，其值在低纳摩尔范围内。小鼠p22HBP的三维结构是通过核磁共振方法确定的，它是SOUL/HBP家族中第一个蛋白质的三维结构，由一个9股扭曲的β桶组成，两侧是两个长α螺旋。尽管在三种细菌蛋白中发现了同源结构域，其中两种是转录因子，但p22HBP确定的折叠对应于真核蛋白中一种新的α+β折叠。化学位移图谱将四吡咯结合位点定位到由αA螺旋和一个延伸环的残基形成的疏水裂缝中。为了评估SOUL/HBP家族中四吡咯结合的结构基础，通过手动对接和自动化方法生成了p22HBP-原卟啉-IX复合物和SOUL蛋白的模型。
