[ASAP] Understanding Which Residues of the Active Site and Loop Structure of a Tyrosine Aminomutase Define Its Mutase and Lyase Activities

May 29th, 2018 by Gayanthi Attanayake, Tyler Walter, Kevin D. Walker

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Biochemistry
DOI: 10.1021/acs.biochem.8b00269

A pathogenesis-related 10 protein catalyzes the final step in thebaine biosynthesis

May 28th, 2018 by Xue Chen

A pathogenesis-related 10 protein catalyzes the final step in thebaine biosynthesis

A pathogenesis-related 10 protein catalyzes the final step in thebaine biosynthesis, Published online: 28 May 2018; doi:10.1038/s41589-018-0059-7

Although the conversion of (7S)-salutaridinol 7-O-acetate to thebaine can occur spontaneously, the identification of a thebaine synthase enzyme that catalyzes the reaction indicates how nature avoids the formation of labile hydroxylated byproducts.
  • Posted in Nat Chem Biol, Publications
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ER-resident protein 46 (ERp46) triggers the mannose-trimming activity of ER degradation-enhancing {alpha}-mannosidase-like protein 3 (EDEM3) [Protein Synthesis and Degradation]

May 21st, 2018 by Shangyu Yu, Shinji Ito, Ikuo Wada, Nobuko Hosokawa

Protein folding in the cell is regulated by several quality-control mechanisms. Correct folding of glycoproteins in the endoplasmic reticulum (ER) is tightly monitored by the recognition of glycan signals by lectins in the ER-associated degradation (ERAD) pathway. In mammals, mannose trimming from N-glycans is crucial for disposal of misfolded glycoproteins. The mannosidases responsible for this process are ER mannosidase I and ER degradation–enhancing α-mannosidase–like proteins (EDEMs). However, the molecular mechanism of mannose removal by EDEMs remains unclear, partly owing to the difficulty of reconstituting mannosidase activity in vitro. Here, our analysis of EDEM3-mediated mannose-trimming activity on a misfolded glycoprotein revealed that ERp46, an ER-resident oxidoreductase, associates stably with EDEM3. This interaction, which depended on the redox activity of ERp46, involved formation of a disulfide bond between the cysteine residues of the ERp46 redox-active sites and the EDEM3 α-mannosidase domain. In a defined in vitro system consisting of recombinant proteins purified from HEK293 cells, the mannose-trimming activity of EDEM3 toward the model misfolded substrate, the glycoprotein T-cell receptor alpha locus (TCRα), was reconstituted only when ERp46 had established a covalent interaction with EDEM3. On the basis of these findings, we propose that disposal of misfolded glycoproteins through mannose trimming is tightly connected to redox-mediated regulation in the ER.
  • Posted in Journal of Biological Chemistry, Publications
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Arginine methylation of translocated in liposarcoma (TLS) inhibits its binding to long noncoding RNA, abrogating TLS-mediated repression of CBP/p300 activity [RNA]

May 21st, 2018 by Wei Cui, Ryoma Yoneda, Naomi Ueda, Riki Kurokawa

Translocated in liposarcoma (TLS) is an RNA-binding protein and a transcription-regulatory sensor of DNA damage. TLS binds promoter-associated noncoding RNA (pncRNA) and inhibits histone acetyltransferase (HAT) activity of CREB-binding protein (CBP)/E1A-binding protein P300 (p300) on the cyclin D1 (CCND1) gene. Although post-translational modifications of TLS, such as arginine methylation, are known to regulate TLS’s nucleocytoplasmic shuttling and assembly in stress granules, its interactions with RNAs remain poorly characterized. Herein, using various biochemical assays, we confirmed the earlier observations that TLS is methylated by protein arginine methyltransferase 1 (PRMT1) in vitro. The arginine methylation of TLS disrupted binding to pncRNA and also prevented binding of TLS to and inhibition of CBP/p300. This result indicated that arginine methylation of TLS abrogates both binding to pncRNA and TLS-mediated inhibition of CBP/p300 HAT activities. We also report that an arginine residue within the Arg–Gly–Gly domain of TLS, Arg-476, serves as the major determinant for binding to pncRNA. Either methylation or mutation of Arg-476 of TLS significantly decreased pncRNA binding and thereby prevented a pncRNA-induced allosteric alteration in TLS that is required for its interaction with CBP/p300. Moreover, unlike wildtype TLS, an R476A TLS mutant did not inhibit CCND1 promoter activity in luciferase reporter assays. Taken together, we propose the hypothesis that arginine methylation of TLS regulates both TLS–nucleic acid and TLS–protein interactions and thereby participates in transcriptional regulation.
  • Posted in Journal of Biological Chemistry, Publications
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Computational redesign of enzymes for regio- and enantioselective hydroamination

May 21st, 2018 by Ruifeng Li

Computational redesign of enzymes for regio- and enantioselective hydroamination

Computational redesign of enzymes for regio- and enantioselective hydroamination, Published online: 21 May 2018; doi:10.1038/s41589-018-0053-0

Computational protein design, without subsequent directed evolution, rapidly provides a set of aspartase variants capable of biocatalytic asymmetric addition of ammonia to substituted acrylates, producing various enantiopure β-amino acids.
  • Posted in Nat Chem Biol, Publications
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Genome-wide mapping reveals that deoxyuridine is enriched in the human centromeric DNA

May 21st, 2018 by Xiaoting Shu

Genome-wide mapping reveals that deoxyuridine is enriched in the human centromeric DNA

Genome-wide mapping reveals that deoxyuridine is enriched in the human centromeric DNA, Published online: 21 May 2018; doi:10.1038/s41589-018-0065-9

A genome-wide uracil profiling technology (termed “dU-seq”), based on selective labeling and biotin pull-down, reveals that uracil is enriched at the centromere of the human genome.
  • Posted in Nat Chem Biol, Publications
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tRNA tracking for direct measurements of protein synthesis kinetics in live cells

May 16th, 2018 by Ivan L. Volkov

tRNA tracking for direct measurements of protein synthesis kinetics in live cells

tRNA tracking for direct measurements of protein synthesis kinetics in live cells, Published online: 16 May 2018; doi:10.1038/s41589-018-0063-y

Combination of single-molecule tracking experiments and machine-learning approaches to monitor diffusional state transitions between ribosome-bound and free tRNAs allows codon resolution measurements of translation kinetics.
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Next-generation biocontainment systems for engineered organisms

May 16th, 2018 by Jeong Wook Lee

Next-generation biocontainment systems for engineered organisms

Next-generation biocontainment systems for engineered organisms, Published online: 16 May 2018; doi:10.1038/s41589-018-0056-x

Synthetic biology offers innovative approaches for engineering biological systems, but also supports the development of biocontainment strategies that ensure the safe application of genetically modified organisms.

Identification of a <i>S. aureus</i> virulence factor by activity-based protein profiling (ABPP)

May 16th, 2018 by Christian S. Lentz

Identification of a S. aureus virulence factor by activity-based protein profiling (ABPP)

Identification of a <i>S. aureus</i> virulence factor by activity-based protein profiling (ABPP), Published online: 16 May 2018; doi:10.1038/s41589-018-0060-1

ABP profiling identifies uncharacterized S. aureus serine hydrolases, including the surface-localized FphB, which processes lipid ester substrates and is required for infection in vivo. An FphB inhibitor reduces in vivo bacterial load.
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Functional phosphorylation

May 16th, 2018 by Mirella Bucci

Functional phosphorylation

Functional phosphorylation, Published online: 16 May 2018; doi:10.1038/s41589-018-0075-7

Functional phosphorylation