Local Conformational Stability of HIV-1 gp120 in Unliganded and CD4-Bound States as Defined by Amide Hydrogen/Deuterium Exchange▿ †

The binding reaction of the HIV-1 gp120 envelope glycoprotein to the CD4 receptor involves exceptional changes in enthalpy and entropy. Crystal structures of gp120 in unliganded and various ligand-bound states, meanwhile, reveal an inner domain able to fold into diverse conformations, a structurally invariant outer domain, and, in the CD4-bound state, a bridging sheet minidomain. These studies, however, provide only hints as to the flexibility of each state. Here we use amide hydrogen/deuterium exchange coupled to mass spectrometry to provide quantifications of local conformational stability for HIV-1 gp120 in unliganded and CD4-bound states. On average, unliganded core gp120 displayed >10,000-fold slower exchange of backbone-amide hydrogens than a theoretically unstructured protein of the same composition, with binding by CD4 reducing the rate of gp120 amide exchange a further 10-fold. For the structurally constant CD4, alterations in exchange correlated well with alterations in binding surface (P value = 0.0004). For the structurally variable gp120, however, reductions in flexibility extended outside the binding surface, and regions of expected high structural diversity (inner domain/bridging sheet) displayed roughly 20-fold more rapid exchange in the unliganded state than regions of low diversity (outer domain). Thus, despite an extraordinary reduction in entropy, neither unliganded gp120 nor free CD4 was substantially unstructured, suggesting that most of the diverse conformations that make up the gp120 unliganded state are reasonably ordered. The results provide a framework for understanding how local conformational stability influences entropic change, conformational diversity, and structural rearrangements in the gp120-CD4 binding reaction

A Common Property of Amyotrophic Lateral Sclerosis-associated Variants: DESTABILIZATION OF THE COPPER/ZINC SUPEROXIDE DISMUTASE ELECTROSTATIC LOOP*

At least 119 mutations in the gene encoding copper/zinc superoxide dismutase (SOD1) cause amyotrophic lateral sclerosis by an unidentified toxic gain of function. We compared the dynamic properties of 13 as-isolated, partially metallated, SOD1 variant enzymes using hydrogen-deuterium exchange. We identified a shared property of these familial amyotrophic lateral sclerosis-related SOD1 variants, namely structural and dynamic change affecting the electrostatic loop (loop VII) of SOD1. Furthermore, SOD1 variants that have severely compromised metal binding affinities demonstrated additional structural and dynamic changes to the zinc-binding loop (loop IV) of SOD1. Although the biological consequences of increased loop VII mobility are not fully understood, this common property is consistent with the hypotheses that SOD1 mutations exert toxicity via aggregation or aberrant association with other cellular constituents