Metagenomic discovery of polybrominated diphenyl ether biosynthesis by marine sponges

March 20th, 2017 by Vinayak Agarwal

Nature Chemical Biology 13, 537 (2017). doi:10.1038/nchembio.2330

Authors: Vinayak Agarwal, Jessica M Blanton, Sheila Podell, Arnaud Taton, Michelle A Schorn, Julia Busch, Zhenjian Lin, Eric W Schmidt, Paul R Jensen, Valerie J Paul, Jason S Biggs, James W Golden, Eric E Allen & Bradley S Moore

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The RecJ2 Protein in the Thermophilic Archaeon Thermoplasma acidophilum Is a 3′ 5′ Exonuclease and Associates with a DNA Replication Complex [Microbiology]

March 16th, 2017 by Hiromi Ogino, Sonoko Ishino, Daisuke Kohda, Yoshizumi Ishino

RecJ/cell division cycle 45 (Cdc45) proteins are widely conserved in the three domains of life, i.e., in Bacteria, Eukarya and Archaea. Bacterial RecJ is a 5′ 3′ exonuclease and functions in DNA repair pathways, while using its 5′ 3′ exonuclease activity. Eukaryotic Cdc45 has no identified enzymatic activity, but participates in the CMG complex so named because it is composed of Cdc45, minichromosome maintenance protein complex (MCM) proteins 2-7, and GINS complex proteins (Sld5, Psf11 to 3). Eukaryotic Cdc45 and bacterial/archaeal RecJ share similar amino acid sequences and are considered functional counterparts. In Archaea, a RecJ homolog in Thermococcus kodakarensis was shown to associate with GINS and accelerate its nuclease activity and was therefore designated GAN (GINS-associated nuclease); however, to date, no archaeal RecJ MCM GINS complex has been isolated. The thermophilic archaeon Thermoplasma acidophilum has two RecJ like proteins, designated TaRecJ1 and TaRecJ2. TaRecJ1 exhibited DNA-specific 5′ 3′exonuclease activity, while TaRecJ2 had 3′ 5′ exonuclease activity and preferred RNA over DNA. TaRecJ2, but not TaRecJ1, formed a stable complex with TaGINS in a 2:1 molar ratio. Furthermore, the TaRecJ2-TaGINS complex stimulated activity of TaMCM helicase in vitro, and the TaRecJ2-TaMCM-TaGINS complex was also observed in vivo. However, TaRecJ2 did not interact with TaMCM directly and was not required for the helicase activation in vitro. These findings suggest that the function of archaeal RecJ in DNA replication evolved divergently from Cdc45 despite conservation of the CMG-like complex formation between Archaea and Eukarya.
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Adjacent channelrhodopsin-2 residues within transmembranes 2 and 7 regulate cation selectivity and distribution of the two open states [Membrane Biology]

March 16th, 2017 by Ryan Richards, Robert E. Dempski

Channelrhodopsin-2 (ChR2) is a light-activated channel that can conduct cations of multiple valencies down the electrochemical gradient. Under continuous light exposure, ChR2 transitions from a high conducting open state (O1) to a low conducting open state (O2) with differing ion selectivity. The molecular basis for the O1 to O2 transition and how ChR2 modulates selectivity between states is currently unresolved. To this end, we used steered molecular dynamics, electrophysiology, and kinetic modeling to identify residues that contribute to gating and selectivity in discrete open states. Analysis of steered molecular dynamics experiments identified three transmembrane residues (V86, K93 and N258) that form a putative barrier to ion translocation. Kinetic modeling of photocurrents generated from ChR2 proteins with conservative mutations at these positions demonstrated that these residues contribute to cation selectivity (V86 and N258), the transition between the two open states (V86), open channel stability, and the hydrogen-bonding network (K93I and K93N). These results suggest that this approach can be used to identify residues that contribute to the open state transitions and the discrete ion selectivity within these states. With the rise of ChR2 use in optogenetics, it will be critical to identify residues that contribute to O1 or O2 selectivity and gating to minimize undesirable effects.
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A Secretion-Amplification Role for Salmonella enterica Translocon Protein SipD

March 16th, 2017 by Anum Azam Glasgow, Han Teng Wong and Danielle Tullman-Ercek

TOC Graphic

ACS Synthetic Biology
DOI: 10.1021/acssynbio.6b00335

A Modular Toolkit for Generating Pichia pastoris Secretion Libraries

March 15th, 2017 by Ulrike Obst, Timothy K. Lu and Volker Sieber

TOC Graphic

ACS Synthetic Biology
DOI: 10.1021/acssynbio.6b00337

Decoding cyclase-dependent assembly of hapalindole and fischerindole alkaloids

March 13th, 2017 by Shasha Li

Nature Chemical Biology 13, 467 (2017). doi:10.1038/nchembio.2327

Authors: Shasha Li, Andrew N Lowell, Sean A Newmister, Fengan Yu, Robert M Williams & David H Sherman

The formation of C–C bonds in an enantioselective fashion to create complex polycyclic scaffolds in the hapalindole- and fischerindole- type alkaloids from Stigonematales cyanobacteria represents a compelling and urgent challenge in adapting microbial biosynthesis as a catalytic platform in drug development. Here we determine the biochemical basis for tri- and tetracyclic core formation in these secondary metabolites, involving a new class of cyclases that catalyze a complex cyclization cascade.

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Structural basis of PROTAC cooperative recognition for selective protein degradation

March 13th, 2017 by Morgan S Gadd

Nature Chemical Biology 13, 514 (2017). doi:10.1038/nchembio.2329

Authors: Morgan S Gadd, Andrea Testa, Xavier Lucas, Kwok-Ho Chan, Wenzhang Chen, Douglas J Lamont, Michael Zengerle & Alessio Ciulli

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The Rrp4–exosome complex recruits and channels substrate RNA by a unique mechanism

March 13th, 2017 by Milos A Cvetkovic

Nature Chemical Biology 13, 522 (2017). doi:10.1038/nchembio.2328

Authors: Milos A Cvetkovic, Jan Philip Wurm, Maxime J Audin, Stefan Schütz & Remco Sprangers

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In silico design of novel probes for the atypical opioid receptor MRGPRX2

March 13th, 2017 by Katherine Lansu

Nature Chemical Biology 13, 529 (2017). doi:10.1038/nchembio.2334

Authors: Katherine Lansu, Joel Karpiak, Jing Liu, Xi-Ping Huang, John D McCorvy, Wesley K Kroeze, Tao Che, Hiroshi Nagase, Frank I Carroll, Jian Jin, Brian K Shoichet & Bryan L Roth

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Two alternative binding mechanisms connect the protein translocation Sec71/Sec72 complex with heat shock proteins [Protein Structure and Folding]

March 12th, 2017 by Arati Tripathi, Elisabet C Mandon, Reid Gilmore, Tom A Rapoport

The biosynthesis of many eukaryotic proteins requires accurate targeting to and translocation across the endoplasmic reticulum (ER) membrane. Post-translational protein translocation in yeast requires both the Sec61 translocation channel, and a complex of four additional proteins: Sec63, Sec62, Sec71, and Sec72. The structure and function of these proteins are largely unknown. This pathway also requires the cytosolic Hsp70 protein Ssa1, but whether Ssa1 associates with the translocation machinery to target protein substrates to the membrane is unclear. Here, we use a combined structural and biochemical approach to explore the role of Sec71/Sec72 subcomplex in post-translational protein translocation. To this end, we report a crystal structure of the Sec71/Sec72 complex, which revealed that Sec72 contains a tetratricopeptide repeat (TPR) domain that is anchored to the ER membrane by Sec71. We also determined the crystal structure of this TPR domain with a C-terminal peptide derived from Ssa1, which suggests how Sec72 interacts with full-length Ssa1. Surprisingly, Ssb1, a cytoplasmic Hsp70 that binds ribosome- associated nascent polypeptide chains also binds to the TPR domain of Sec72, even though it lacks the TPR-binding C-terminal residues of Ssa1. We demonstrate that Ssb1 binds through its ATPase domain to the TPR domain, an interaction that leads to inhibition of nucleotide exchange. Taken together, our results suggest that translocation substrates can be recruited to the Sec71/72 complex either post-translationally through Ssa1 or co-translationally through Ssb1.
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