Kavanaugh Jeffrey S, Rogers Paul H, Arnone Arthur
Department of Biochemistry, Roy J. and Lucille A. Carver College of Medicine, The University of Iowa, Iowa City, Iowa 52242, USA.
Biochemistry. 2005 Apr 26;44(16):6101-21. doi: 10.1021/bi047813a.
A detailed description of hemoglobin cooperativity requires knowledge of the dimer-dimer interactions responsible for the low ligand affinity of the quaternary-T tetramer, the "quaternary-T constraints", along with stereochemical pathways that specify how ligand binding disrupts these quaternary constraints. The recent mutagenic screen of Noble et al. [Noble, R. W., et al. (2001) Biochemistry 40, 12357-12368] has identified the major region of quaternary constraint to be a cluster of residues at the alpha1beta2 interface that is centered at Trp37beta. In this paper, crystallographic studies are presented for most of the mutant hemoglobins studied by Noble et al. These crystallographic experiments identify structural transitions-referred to as T-to-T(High) transitions-between the quaternary-T structure of wild-type deoxyhemoglobin and an ensemble of related T-like quaternary structures that are induced by some mutations in the Trp37beta cluster and/or by exposing crystals of wild-type or mutant deoxyhemoglobins to oxygen. The T-to-T(High) quaternary transitions consist of a rotation of the alpha1beta1 dimer relative to the alpha2beta2 dimer as well as a coupled alphabeta dimer bending component that consists of a small rotation of the alpha1 subunit relative to the beta1 subunit (and a symmetry related rotation of the alpha2 subunit relative to the beta2 subunit). In addition, differences in subunit tertiary structure associated with the T-to-T(High) transitions suggest two stereochemical pathways (one associated with the alpha subunits and one associated with the betasubunits) by which ligand binding specifically disrupts quaternary constraints in the Trp37beta cluster. In the alpha subunits, ligand binding induces a shift of the heme iron producing tension in a chain of covalent bonds that extends from the Fe-N(epsilon)(2)His(F8)alpha1 bond to the peptide backbone bonds of residues His87(F8)alpha1 and Ala88(F9)alpha1. This tension induces an alpha-to-pi transition in the COOH-terminal end of the F-helix that shifts the beta-carbon of Ala88alpha1 by approximately 1.5 A directly into the side chain of Tyr140alpha1 (a key residue in the Trp37beta2 cluster). Collectively these structural changes constitute a relatively short pathway by which ligand binding forces Tyr140alpha1 into the alpha1beta2 interface disrupting quaternary constraints associated with the Trp37beta2 cluster. In the beta subunits, our analysis suggests a more extended energy transduction pathway in which ligand-induced beta1-heme movement triggers tertiary changes in the beta1 subunit that promote alpha1beta1 dimer bending that disrupts quaternary constraints in the Trp37beta2 cluster at the alpha1beta2 interface.
对血红蛋白协同性的详细描述需要了解导致四级结构T型四聚体配体亲和力较低的二聚体-二聚体相互作用，即“四级结构T型限制”，以及规定配体结合如何破坏这些四级限制的立体化学途径。Noble等人最近的诱变筛选[Noble, R. W., 等人 (2001) Biochemistry 40, 12357 - 12368]已确定四级限制的主要区域是α1β2界面处的一组残基，该区域以β链的Trp37为中心。本文展示了Noble等人研究的大多数突变血红蛋白的晶体学研究。这些晶体学实验确定了野生型脱氧血红蛋白的四级结构T型与一组相关的T样四级结构之间的结构转变——称为T型到T(高)型转变，这些转变是由Trp37β簇中的一些突变和/或通过将野生型或突变型脱氧血红蛋白晶体暴露于氧气中诱导产生的。T型到T(高)型的四级转变包括α1β1二聚体相对于α2β2二聚体的旋转以及一个耦合的αβ二聚体弯曲成分，该成分由α1亚基相对于β1亚基的小旋转（以及α2亚基相对于β2亚基的对称相关旋转）组成。此外，与T型到T(高)型转变相关的亚基三级结构差异表明了两条立体化学途径（一条与α亚基相关，一条与β亚基相关），通过这些途径配体结合特异性地破坏了Trp37β簇中的四级限制。在α亚基中，配体结合会导致血红素铁的移动，在从Fe-N(ε)(2)His(F8)α1键延伸到His87(F8)α1和Ala88(F9)α1残基的肽主链键的共价键链中产生张力。这种张力在F-螺旋的COOH末端诱导α到π的转变，使Ala88α1的β-碳直接向Tyr140α1（Trp37β2簇中的一个关键残基）的侧链移动约1.5 Å。这些结构变化共同构成了一条相对较短的途径，通过该途径配体结合将Tyr140α1强行拉入α1β2界面，破坏了与Trp37β2簇相关的四级限制。在β亚基中，我们的分析表明存在一条更长的能量转导途径，其中配体诱导的β1-血红素移动触发β1亚基的三级变化，促进α1β1二聚体弯曲，从而破坏α1β2界面处Trp37β2簇中的四级限制。
