All-atom molecular dynamics comparison of disease-associated zinc fingers

<p>An important regulatory domain of NF-B Essential Modulator (NEMO) is a ubiquitin-binding zinc finger, with a tetrahedral CYS3HIS1 zinc-coordinating binding site. Two variations of NEMO’s zinc finger are implicated in various disease states including ectodermal dysplasia and adult-onset glaucoma. To discern structural and dynamical differences between these disease states, we present results of 48-s of molecular dynamics simulations for three zinc finger systems each in two states, with and without zinc-bound and correspondingly appropriate cysteine thiol/thiolate configurations. The wild-type protein, often studied for its role in cancer, maintains the most rigid and conformationally stable zinc-bound configuration compared with the diseased counterparts. The glaucoma-related protein has persistent loss of secondary structure except within the dominant conformation. Conformational overlap between wild-type and glaucoma isoforms indicate a competitive binding mechanism may be substantial in the malfunctioning configuration, while the alpha-helical disruption of the ectodermal dysplasia suggests a loss of binding selectivity is responsible for aberrant function.</p

Binding Site Configurations Probe the Structure and Dynamics of the Zinc Finger of NEMO (NF-κB Essential Modulator)

Zinc-finger proteins are regulators of critical signaling pathways for various cellular functions, including apoptosis and oncogenesis. Here, we investigate how binding site protonation states and zinc coordination influence protein structure, dynamics, and ultimately function, as these pivotal regulatory proteins are increasingly important for protein engineering and therapeutic discovery. To better understand the thermodynamics and dynamics of the zinc finger of NEMO (NF-κB essential modulator), as well as the role of zinc, we present results of 20 μs molecular dynamics trajectories, 5 μs for each of four active site configurations. Consistent with experimental evidence, the zinc ion is essential for mechanical stabilization of the functional, folded conformation. Hydrogen bond motifs are unique for deprotonated configurations yet overlap in protonated cases. Correlated motions and principal component analysis corroborate the similarity of the protonated configurations and highlight unique relationships of the zinc-bound configuration. We hypothesize a potential mechanism for zinc binding from results of the thiol configurations. The deprotonated, zinc-bound configuration alone predominantly maintains its tertiary structure throughout all 5 μs and alludes rare conformations potentially important for (im)­proper zinc-finger-related protein–protein or protein–DNA interactions

Uncovering Large-Scale Conformational Change in Molecular Dynamics without Prior Knowledge

As the length of molecular dynamics (MD) trajectories grows with increasing computational power, so does the importance of clustering methods for partitioning trajectories into conformational bins. Of the methods available, the vast majority require users to either have some <i>a priori</i> knowledge about the system to be clustered or to tune clustering parameters through trial and error. Here we present non-parametric uses of two modern clustering techniques suitable for first-pass investigation of an MD trajectory. Being non-parametric, these methods require neither prior knowledge nor parameter tuning. The first method, HDBSCAN, is fastrelative to other popular clustering methodsand is able to group unstructured or intrinsically disordered systems (such as intrinsically disordered proteins, or IDPs) into bins that represent global conformational shifts. HDBSCAN is also useful for determining the overall stability of a systemas it tends to group stable systems into one or two binsand identifying transition events between metastable states. The second method, iMWK-Means, with explicit rescaling followed by K-Means, while slower than HDBSCAN, performs well with stable, structured systems such as folded proteins and is able to identify higher resolution details such as changes in relative position of secondary structural elements. Used in conjunction, these clustering methods allow a user to discern quickly and without prior knowledge the stability of a simulated system and identify both local and global conformational changes