Maybe Just a Little Bit Special, After All?

The attitude—common among tax professionals—that tax is special (mostly because of its supposedly unique complexity), and that special legal rules should apply in the tax context, has been described and excoriated by scholars as tax exceptionalism or tax myopia. The Supreme Court dealt tax exceptionalism a grievous blow in its 2011 opinion in Mayo Foundation for Medical Education & Research v. United States, in which it held that the Chevron standard for determining the validity of regulations applied in tax just as it applied in other fields. One commentator gleefully celebrated Mayo as the death knell of tax exceptionalism, declaring, The tax world finally recognized a stark fact of life in 2011: Tax law is not special. This Article offers, with numerous hedges and qualifications, a defense of the exceptionalists and of exceptionalism. It makes three points for the defense. First, it is not so much tax professionals who think tax is special; rather, the view of tax as a thing apart is held most strongly by everyone else. Second, to the extent tax professionals do believe that tax is special, they resemble antitrust lawyers who think that antitrust is special, bankruptcy lawyers who think that bankruptcy is special, and so on. In other words, there is nothing exceptional about tax exceptionalism. And, finally, to the extent tax professionals not only think tax is special but also think it is more special than, say, antitrust lawyers think that antitrust is special, they may not be altogether wrong. Maybe tax really is just a little bit special, after all

Folding and Misfolding in a Naturally Occurring Circularly Permuted PDZ Domain

One of the most extreme and fascinating examples of naturally occurring mutagenesis is represented by circular permutation. Circular permutations involve the linking of two chain ends and cleavage at another site. Here we report the first description of the folding mechanism of a naturally occurring circularly permuted protein, a PDZ domain from the green alga Scenedesmus obliquus. Data reveal that the folding of the permuted protein is characterized by the presence of a low energy off-pathway kinetic trap. This finding contrasts with what was previously observed for canonical PDZ domains that, although displaying a similar primary structure when structurally re-aligned, fold via an on-pathway productive intermediate. Although circular permutation of PDZ domains may be necessary for a correct orientation of their functional sites in multi-domain protein scaffolds, such structural rearrangement may compromise their folding pathway. This study provides a straightforward example of the divergent demands of folding and function

Proteomic peptide phage display uncovers novel interactions of the PDZ1-2 supramodule of syntenin

Syntenin has crucial roles in cell adhesion, cell migration and synaptic transmission. Its closely linked postsynaptic density-95, discs large 1, zonula occludens-1 (PDZ) domains typically interact with C-terminal ligands. We profile syntenin PDZ1-2 through proteomic peptide phage display (ProP-PD) using a library that displays C-terminal regions of the human proteome. The protein recognizes a broad range of peptides, with a preference for hydrophobic motifs and has a tendency to recognize cryptic internal ligands. We validate the interaction with nectin-1 through orthogonal assays. The study demonstrates the power of ProP-PD as a complementary approach to uncover interactions of potential biological relevance

Large-scale interaction profiling of PDZ domains through proteomic peptide-phage display using human and viral phage peptidomes

The human proteome contains a plethora of short linear motifs (SLiMs) that serve as binding interfaces for modular protein domains. Such interactions are crucial for signaling and other cellular processes, but are difficult to detect because of their low to moderate affinities. Here we developed a dedicated approach, proteomic peptide-phage display (ProP-PD), to identify domain-SLiM interactions. Specifically, we generated phage libraries containing all human and viral C-terminal peptides using custom oligonucleotide microarrays. With these libraries we screened the nine PSD-95/ Dlg/ZO-1 (PDZ) domains of human Densin-180, Erbin, Scribble, and Disks large homolog 1 for peptide ligands. We identified several known and putative interactions potentially relevant to cellular signaling pathways and confirmed interactions between fulllength Scribble and the target proteins β-PIX, plakophilin-4, and guanylate cyclase soluble subunit a-2 using colocalization and coimmunoprecipitation experiments. The affinities of recombinant Scribble PDZ domains and the synthetic peptides representing the C termini of these proteins were in the 1- to 40-μM range. Furthermore, we identified several well-established host-virus protein- protein interactions, and confirmed that PDZ domains of Scribble interact with the C terminus of Tax-1 of human T-cell leukemia virus with micromolar affinity. Previously unknown putative viral protein ligands for the PDZ domains of Scribble and Erbin were also identified. Thus, we demonstrate that our ProP-PD libraries are useful tools for probing PDZ domain interactions. The method can be extended to interrogate all potential eukaryotic, bacterial, and viral SLiMs and we suggest it will be a highly valuable approach for studying cellular and pathogen-host protein-protein interactions

Evolutionary Analysis and Posttranslational Chemical Modifications in Protein Redesign : A Study on Mu Class Glutathione Transferases

Glutathione transferases (GSTs) constitute a family of multifarious enzymes that conjugate glutathione (GSH) with a wide range of electrophiles. GSTs are grouped into different classes based on protein sequence similarities. Despite high sequence identities between GSTs of the same class they often display different substrate specificites. Human GST M1-1 is efficiently catalyzing the conjugation of GSH and various epoxide substrates, whereas the 84% sequence-identical GST M2-2 has low activities with the same substrates. Evolutionary rate analysis was used to identify hypervariable amino acid positions among GST Mu class sequences. A Thr to Ser conversion of the variable residue 210 in GST M2-2 elicited a drastic increase in catalytic activity with epoxides, which is the characteristic activity of GST M1-1. This provides support for the usefulness of evolutionary analysis in identifying functionally important residues, although the additional mutations of two other variable residues did not confer any noteworthy changes in activity. To further investigate the functional importance of residue T210 in GST M2-2 it was replaced by all other commonly occurring amino acids. The replacements caused marked changes in substrate specificity, stability, and expressivity, indicating how functionalities of a duplicated Mu class GST may easily be altered by point mutations. The stereo- and regioselectivity in epoxide-conjugation catalyzed by GSTs M1-1 and M2-2 was investigated. The results show that a serine in position 210 is beneficial for high enantioselectivity with trans-stilbene oxide. However, an alanine in position 210 is more favorable for stereo- and regioselectivity with the smaller epoxide substrate styrene-7,8-oxide. The low enantioselectivity of GST M1-1 was improved 10- and 9- fold with styrene-7,8-oxide and 1-phenylpropylene oxide, respectively, through different combination of site-specific mutations and posttranslational chemical modifications. The approach can be employed in more extensive screening experiments where a large variety of modifications easily can be tested

High-throughput methods for identification of protein-protein interactions involving short linear motifs

Interactions between modular domains and short linear motifs (3-10 amino acids peptide stretches) are crucial for cell signaling. The motifs typically reside in the disordered regions of the proteome and the interactions are often transient, allowing for rapid changes in response to changing stimuli. The properties that make domain-motif interactions suitable for cell signaling also make them difficult to capture experimentally and they are therefore largely underrepresented in the known protein-protein interaction networks. Most of the knowledge on domain-motif interactions is derived from low-throughput studies, although there exist dedicated high-throughput methods for the identification of domain-motif interactions. The methods include arrays of peptides or proteins, display of peptides on phage or yeast, and yeast-two-hybrid experiments. We here provide a survey of scalable methods for domain-motif interaction profiling. These methods have frequently been applied to a limited number of ubiquitous domain families. It is now time to apply them to a broader set of peptide binding proteins, to provide a comprehensive picture of the linear motifs in the human proteome and to link them to their potential binding partners. Despite the plethora of methods, it is still a challenge for most approaches to identify interactions that rely on post-translational modification or context dependent or conditional interactions, suggesting directions for further method development

Affinity and specificity of motif-based protein-protein interactions

It is becoming increasingly clear that eukaryotic cell physiology is largely controlled by protein protein interactions involving disordered protein regions, which usually interact with globular domains in a coupled binding and folding reaction. Several protein recognition domains are part of large families where members can interact with similar peptide ligands. Because of this, much research has been devoted to understanding how specificity can be achieved. A combination of interface complementarity, interactions outside of the core binding site, avidity from multidomain architecture and spatial and temporal regulation of expression resolves the conundrum. Here, we review recent advances in molecular aspects of affinity and specificity in such protein-protein interactions

Regio- and enantioselectivities in epoxide conjugations are modulated by residue 210 in Mu class glutathione transferases

The homologous human glutathione transferases (GSTs) M1-1 and M2-2 have similar catalytic activities with many electrophilic substrates, but differ strikingly in their conjugation of epoxides with glutathione. Residue 210, Thr in GST M2-2 and Ser in GST M1-1, is a key active-site component in determining the activity profile with epoxide substrates. This residue is hypervariable in Mu class GSTs, suggesting that it has special significance in the evolution of new functions. The present study shows that minor modifications of this residue can have major consequences for the enzyme-catalyzed epoxide conjugations. In general, a Ser at position 210 gives the highest catalytic efficiency, but the relatively high activity with an Ala placed on this position demonstrates that a hydroxyl group is not required. In contrast, a Thr residue suppresses the activity with epoxides by several orders of magnitude without major effects on the activity with alternative GST substrates. Residue 210 influences both the regio- and enantioselectivity with chiral and prochiral epoxides of stilbene and styrene and influences the distribution of isomeric glutathione conjugates. Thus, residue 210 contributes to both stereoselective recognition of the substrates and to partitioning of the isomeric reactants to the alternative transition states leading to separate chiral products.status: publishe

Genetically Encoded Cyclic Peptide Phage Display Libraries

Phage display of self-cycling peptides as an efficient and scalable approach to discover macrocyclic inhibitors of protein-protein interactions