Somatic mutations in PI3Kα: Structural basis for enzyme activation and drug design

The immune response to phosphocholine (PC)–protein is characterized by a shift in antibody repertoire as the response progresses. This change in expressed gene combinations is accompanied by a shift in fine specificity toward the carrier, resulting in high affinity to PC–protein. The somatically mutated memory hybridoma, M3C65, possesses high affinity for PC–protein and the phenyl-hapten analogue, p-nitrophenyl phosphocholine (NPPC). Affinity measurements using related PC–phenyl analogues, including peptides of varying lengths, demonstrate that carrier determinants contribute to binding affinity and that somatic mutations alter this recognition. The crystal structure of an M3C65–NPPC complex at 2.35-Å resolution allows evaluation of the three light chain mutations that confer high-affinity binding to NPPC. Only one of the mutations involves a contact residue, whereas the other two have indirect effects on the shape of the combining site. Comparison of the M3C65 structure to that of T15, an antibody dominating the primary response, provides clear structural evidence for the role of carrier determinants in promoting repertoire shift. These two antibodies express unrelated variable region heavy and light chain genes and represent a classic example of the effect of repertoire shift on maturation of the immune response.

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Hepatitis B Virus Capsids Have Diverse Structural Responses to Small-Molecule Ligands Bound to the Heteroaryldihydropyrimidine Pocket

Journal of Virology ◽

10.1128/jvi.03058-15 ◽

2016 ◽

Vol 90 (8) ◽

pp. 3994-4004 ◽

Cited By ~ 40

Author(s):

Balasubramanian Venkatakrishnan ◽

Sarah P. Katen ◽

Samson Francis ◽

Srinivas Chirapu ◽

M. G. Finn ◽

...

Keyword(s):

Life Cycle ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Drug Design ◽

Quaternary Structure ◽

Core Protein ◽

Viral Life Cycle ◽

Structural Basis ◽

Allosteric Modulators ◽

B Virus

ABSTRACTThough the hepatitis B virus (HBV) core protein is an important participant in many aspects of the viral life cycle, its best-characterized activity is self-assembly into 240-monomer capsids. Small molecules that target core protein (core protein allosteric modulators [CpAMs]) represent a promising antiviral strategy. To better understand the structural basis of the CpAM mechanism, we determined the crystal structure of the HBV capsid in complex with HAP18. HAP18 accelerates assembly, increases protein-protein association more than 100-fold, and induces assembly of nonicosahedral macrostructures. In a preformed capsid, HAP18 is found at quasiequivalent subunit-subunit interfaces. In a detailed comparison to the two other extant CpAM structures, we find that the HAP18-capsid structure presents a paradox. Whereas the two other structures expanded the capsid diameter by up to 10 Å, HAP18 caused only minor changes in quaternary structure and actually decreased the capsid diameter by ∼3 Å. These results indicate that CpAMs do not have a single allosteric effect on capsid structure. We suggest that HBV capsids present an ensemble of states that can be trapped by CpAMs, indicating a more complex basis for antiviral drug design.IMPORTANCEHepatitis B virus core protein has multiple roles in the viral life cycle—assembly, compartment for reverse transcription, intracellular trafficking, and nuclear functions—making it an attractive antiviral target. Core protein allosteric modulators (CpAMs) are an experimental class of antivirals that bind core protein. The most recognized CpAM activity is that they accelerate core protein assembly and strengthen interactions between subunits. In this study, we observe that the CpAM-binding pocket has multiple conformations. We compare structures of capsids cocrystallized with different CpAMs and find that they also affect quaternary structure in different ways. These results suggest that the capsid “breathes” and is trapped in different states by the drug and crystallization. Understanding that the capsid is a moving target will aid drug design and improve our understanding of HBV interaction with its environment.

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dUTPase as a Platform for Antimalarial Drug Design: Structural Basis for the Selectivity of a Class of Nucleoside Inhibitors

Structure ◽

10.1016/j.str.2004.11.015 ◽

2005 ◽

Vol 13 (2) ◽

pp. 329-338 ◽

Cited By ~ 65

Author(s):

Jean L. Whittingham ◽

Isabel Leal ◽

Corinne Nguyen ◽

Ganasan Kasinathan ◽

Emma Bell ◽

...

Keyword(s):

Drug Design ◽

Antimalarial Drug ◽

Structural Basis ◽

Nucleoside Inhibitors

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Lipid peroxidation and phospholipase A2 activity in liposomes composed of unsaturated phospholipids: a structural basis for enzyme activation

Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism ◽

10.1016/0005-2760(88)90079-3 ◽

1988 ◽

Vol 961 (3) ◽

pp. 316-327 ◽

Cited By ~ 116

Author(s):

Alex Sevanian ◽

Mary Lou Wratten ◽

Laurie L. McLeod ◽

Eunjoo Kim

Keyword(s):

Lipid Peroxidation ◽

Phospholipase A2 ◽

Enzyme Activation ◽

Structural Basis ◽

Phospholipase A2 Activity

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Small-molecule crystal structures as a structural basis for drug design

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s0907444994014526 ◽

1995 ◽

Vol 51 (4) ◽

pp. 407-417 ◽

Cited By ~ 12

Author(s):

C. Pascard

Keyword(s):

Drug Design ◽

Crystal Structures ◽

Small Molecule ◽

Structural Basis

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Structure-Based Drug Design of Novel Aurora Kinase A Inhibitors: Structural Basis for Potency and Specificity

Journal of Medicinal Chemistry ◽

10.1021/jm801270e ◽

2009 ◽

Vol 52 (4) ◽

pp. 1050-1062 ◽

Cited By ~ 51

Author(s):

Mohane Selvaraj Coumar ◽

Jiun-Shyang Leou ◽

Paritosh Shukla ◽

Jian-Sung Wu ◽

Ajay Kumar Dixit ◽

...

Keyword(s):

Drug Design ◽

Aurora Kinase ◽

Structural Basis ◽

Structure Based Drug Design ◽

Aurora Kinase A ◽

Kinase A

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Structural basis for the RNA-guided ribonuclease activity of CRISPR-Cas13d

10.1101/314401 ◽

2018 ◽

Author(s):

Cheng Zhang ◽

Silvana Konermann ◽

Nicholas J. Brideau ◽

Peter Lotfy ◽

Scott J. Novick ◽

...

Keyword(s):

Conformational Dynamics ◽

Enzyme Activation ◽

Structural Basis ◽

Binary Complex ◽

Molecular Architecture ◽

Guide Rna ◽

Guide Rnas ◽

Rna Targeting ◽

Hydrogen Deuterium ◽

Target Rna

AbstractCRISPR-Cas endonucleases directed against foreign nucleic acids mediate prokaryotic adaptive immunity and have been tailored for broad genetic engineering applications. Type VI-D CRISPR systems contain the smallest known family of single effector Cas enzymes, and their signature Cas13d ribonuclease employs guide RNAs to cleave matching target RNAs. To understand the molecular basis for Cas13d function, we resolved cryo-electron microscopy structures of Cas13d-guide RNA binary complex and Cas13d-guide-target RNA ternary complex to 3.4 and 3.3 Å resolution, respectively. Furthermore, a 6.5 Å reconstruction of apo Cas13d combined with hydrogen-deuterium exchange revealed conformational dynamics that have implications for RNA scanning. These structures, together with biochemical and cellular characterization, explain the compact molecular architecture of Cas13d and provide insights into the structural transitions required for enzyme activation. Our comprehensive analysis of Cas13d in diverse enzymatic states facilitated site-specific truncations for minimal size and delineates a blueprint for improving biomolecular applications of RNA targeting.

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