Allosteric inhibition of PPM1D serine/threonine phosphatase via an altered conformational state.

PPM1D encodes a serine/threonine phosphatase that regulates numerous pathways including the DNA damage response and p53. Activating mutations and amplification of PPM1D are found across numerous cancer types. GSK2830371 is a potent and selective allosteric inhibitor of PPM1D, but its mechanism of binding and inhibition of catalytic activity are unknown.

Activation of retinal ganglion cells using a biomimetic artificial retina.

Biomimetic protein-based artificial retinas offer a new paradigm for restoring vision for patients blinded by retinal degeneration. Artificial retinas, comprised of an ion-permeable membrane and alternating layers of bacteriorhodopsin (BR) and a polycation binder, are assembled using layer-by-layer electrostatic adsorption. Upon light absorption, the oriented BR layers generate a unidirectional proton gradient.

Molecular basis for substrate recruitment to the PRMT5 methylosome.

PRMT5 is an essential arginine methyltransferase and a therapeutic target in MTAP-null cancers. PRMT5 uses adaptor proteins for substrate recruitment through a previously undefined mechanism. Here, we identify an evolutionarily conserved peptide sequence shared among the three known substrate adaptors (CLNS1A, RIOK1, and COPR5) and show that it is necessary and sufficient for interaction with PRMT5.

Discovery of a First-in-Class Inhibitor of the PRMT5-Substrate Adaptor Interaction.

PRMT5 and its substrate adaptor proteins (SAPs), pICln and Riok1, are synthetic lethal dependencies in MTAP-deleted cancer cells. SAPs share a conserved PRMT5 binding motif (PBM) which mediates binding to a surface of PRMT5 distal to the catalytic site. This interaction is required for methylation of several PRMT5 substrates, including histone and spliceosome complexes.

Structure of PDE3A-SLFN12 complex reveals requirements for activation of SLFN12 RNase.

DNMDP and related compounds, or velcrins, induce complex formation between the phosphodiesterase PDE3A and the SLFN12 protein, leading to a cytotoxic response in cancer cells that express elevated levels of both proteins. The mechanisms by which velcrins induce complex formation, and how the PDE3A-SLFN12 complex causes cancer cell death, are not fully understood. Here, we show that PDE3A and SLFN12 form a heterotetramer stabilized by binding of DNMDP.

Assessing optimal: inequalities in codon optimization algorithms.

Background: Custom genes have become a common resource in recombinant biology over the last 20 years due to the plummeting cost of DNA synthesis. These genes are often "optimized" to non-native sequences for overexpression in a non-native host by substituting synonymous codons within the coding DNA sequence (CDS). A handful of studies have compared native and optimized CDSs, reporting different levels of soluble product due to the accumulation of misfolded aggregates, variable activity of enzymes, and (at least one report of) a change in substrate specificity.

High-Throughput Screens To Identify Autophagy Inducers That Function by Disrupting Beclin 1/Bcl-2 Binding.

Autophagy, a lysosomal degradation pathway, plays a crucial role in cellular homeostasis, development, immunity, tumor suppression, metabolism, prevention of neurodegeneration, and lifespan extension. Thus, pharmacological stimulation of autophagy may be an effective approach for preventing or treating certain human diseases and/or aging. We sought to establish a method for developing new chemical compounds that specifically induce autophagy.

The Autophagy-Related Beclin-1 Protein Requires the Coiled-Coil and BARA Domains To Form a Homodimer with Submicromolar Affinity.

Beclin-1 (BECN1) is an essential component of macroautophagy. This process is a highly conserved survival mechanism that recycles damaged cellular components or pathogens by encasing them in a bilayer vesicle that fuses with a lysosome to allow degradation of the vesicular contents. Mutations or altered expression profiles of BECN1 have been linked to various cancers and neurodegenerative diseases.

Proteolytic control of the mitochondrial calcium uniporter complex.

The mitochondrial calcium uniporter is a Ca-activated Ca channel complex mediating mitochondrial Ca uptake, a process crucial for Ca signaling, bioenergetics, and cell death. The uniporter is composed of the pore-forming MCU protein, the gatekeeping MICU1 and MICU2 subunits, and EMRE, a single-pass membrane protein that links MCU and MICU1 together. As a bridging subunit required for channel function, EMRE could paradoxically inhibit uniporter complex formation if expressed in excess.

Dual functions of a small regulatory subunit in the mitochondrial calcium uniporter complex.

Mitochondrial Ca(2+) uptake, a process crucial for bioenergetics and Ca(2+) signaling, is catalyzed by the mitochondrial calcium uniporter. The uniporter is a multi-subunit Ca(2+)-activated Ca(2+) channel, with the Ca(2+) pore formed by the MCU protein and Ca(2+)-dependent activation mediated by MICU subunits. Recently, a mitochondrial inner membrane protein EMRE was identified as a uniporter subunit absolutely required for Ca(2+) permeation.

Crystal Structure of Recoverin with Calcium Ions Bound to Both Functional EF Hands.

Recoverin (Rv), a small Ca(2+)-binding protein that inhibits rhodopsin kinase (RK), has four EF hands, two of which are functional (EF2 and EF3). Activation requires Ca(2+) in both EF hands, but crystal structures have never been observed with Ca(2+) ions in both sites; all previous structures have Ca(2+) bound to only EF3. We suspected that this was due to an intermolecular crystal contact between T80 and a surface glutamate (E153) that precluded coordination of a Ca(2+) ion in EF2.

Photochromic bacteriorhodopsin mutant with high holographic efficiency and enhanced stability via a putative self-repair mechanism.

The Q photoproduct of bacteriorhodopsin (BR) is the basis of several biophotonic technologies that employ BR as the photoactive element. Several blue BR (bBR) mutants, generated by using directed evolution, were investigated with respect to the photochemical formation of the Q state. We report here a new bBR mutant, D85E/D96Q, which is capable of efficiently converting the entire sample to and from the Q photoproduct.

A highly conserved cysteine of neuronal calcium-sensing proteins controls cooperative binding of Ca2+ to recoverin.

Recoverin, a 23-kDa Ca(2+)-binding protein of the neuronal calcium sensing (NCS) family, inhibits rhodopsin kinase, a Ser/Thr kinase responsible for termination of photoactivated rhodopsin in rod photoreceptor cells. Recoverin has two functional EF hands and a myristoylated N terminus. The myristoyl chain imparts cooperativity to the Ca(2+)-binding sites through an allosteric mechanism involving a conformational equilibrium between R and T states of the protein.

Directed evolution of bacteriorhodopsin for applications in bioelectronics.

In nature, biological systems gradually evolve through complex, algorithmic processes involving mutation and differential selection. Evolution has optimized biological macromolecules for a variety of functions to provide a comparative advantage. However, nature does not optimize molecules for use in human-made devices, as it would gain no survival advantage in such cooperation.

meso-arylporpholactones and their reduction products.

The rational syntheses of meso-tetraaryl-3-oxo-2-oxaporphyrins 5, known as porpholactones, via MnO(4)(-)-mediated oxidations of the corresponding meso-tetraaryl-2,3-dihydroxychlorins (7) is detailed. Since chlorin 7 is prepared from the parent porphyrin 1, this amounts to a 2-step replacement of a pyrrole moiety in 1 by an oxazolone moiety. The stepwise reduction of the porpholactone 5 results in the formation of chlorin analogues, meso-tetraaryl-3-hydroxy-2-oxachlorin (11) and meso-tetraaryl-2-oxachlorins (12).

Green proteorhodopsin reconstituted into nanoscale phospholipid bilayers (nanodiscs) as photoactive monomers.

Over 4000 putative proteorhodopsins (PRs) have been identified throughout the oceans and seas of the Earth. The first of these eubacterial rhodopsins was discovered in 2000 and has expanded the family of microbial proton pumps to all three domains of life. With photophysical properties similar to those of bacteriorhodopsin, an archaeal proton pump, PRs are also generating interest for their potential use in various photonic applications.

Photochemical and thermal stability of green and blue proteorhodopsins: implications for protein-based bioelectronic devices.

The photochemical and thermal stability of the detergent-solubilized blue- and green-absorbing proteorhodpsins, BPR and GPR, respectively, are investigated to determine the viability of these proteins for photonic device applications. Photochemical stability is studied by using pulsed laser excitation and differential UV-vis spectroscopy to assign the photocyclicity. GPR, with a cyclicity of 7 × 10(4) photocycles protein(-1), is 4-5 times more stable than BPR (9 × 10(3) photocycles protein(-1)), but is less stable than native bacteriorhodopsin (9 × 10(5) photocycles protein(-1)) or the 4-keto-bacteriorhodopsin analogue (1 × 10(5) photocycles protein(-1)).

A second-site suppressor of a folding defect functions via interactions with a chaperone network to improve folding and assembly in vivo.

Single amino acid substitutions in a protein can cause misfolding and aggregation to occur. Protein misfolding can be rescued by second-site amino acid substitutions called suppressor substitutions (su), commonly through stabilizing the native state of the protein or by increasing the rate of folding. Here we report evidence that su substitutions that rescue bacteriophage P22 temperature-sensitive-folding (tsf) coat protein variants function in a novel way.