scholarly journals Computational and experimental assessment of backbone templates for computational protein design

2021 ◽  
Author(s):  
Frederikke I Marin ◽  
Kristoffer E Johansson ◽  
Charlotte O'Shea ◽  
Kresten Lindorff-Larsen ◽  
Jakob R Winther
Keyword(s):  
Protein Design ◽  
Screening Tool ◽  
Biochemical Characterization ◽  
State Of The Art ◽  
Computational Protein Design ◽  
Energy Term ◽  
E Coli ◽  
Design Software ◽  
Folded Structures ◽  
At Will

Computational protein design has taken big strides over the recent years, however, the tools available are still not at a state where a sequence can be designed to fold into a given protein structure at will and with high probability. We have here applied a recent release of Rosetta Design to redesign a set of structurally very similar proteins belonging to the Thioredoxin fold. We determined design success using a combination of a genetic screening tool to assay folding/stability in E. coli and selecting the best hits from this for further biochemical characterization. We have previously used this set of template proteins for redesign and found that success was highly dependent on template structure, a trait which was also found in this study. Nevertheless, state of the art design software is now able to predict the best template, most likely due to the introduction of the cart_bonded energy term. The template that led to the greatest fraction of successful designs was the same (a Thioredoxin from spinach) as that identified in our previous study. Our previously described redesign of Thioredoxin, which also used the spinach protein as template, however also performed well. In the present study, both these templates yielded proteins with compact folded structures, and enforces the conclusion that any design project must carefully consider different design templates. Fortunately, selecting designs using the cart_bonded energy term appears to correctly identify such templates.

scholarly journals cOSPREY: A Cloud-Based Distributed Algorithm for Large-Scale Computational Protein Design

2016 ◽  
Vol 23 (9) ◽  
pp. 737-749 ◽  
Author(s):  
Yuchao Pan ◽  
Yuxi Dong ◽  
Jingtian Zhou ◽  
Mark Hallen ◽  
Bruce R. Donald ◽  
...  
Keyword(s):  
Protein Design ◽  
Distributed Algorithm ◽  
Large Scale ◽  
Computational Protein Design

scholarly journals Discovery of Substrates for a SET Domain Lysine Methyltransferase Predicted by Multistate Computational Protein Design

Structure ◽  
2015 ◽  
Vol 23 (1) ◽  
pp. 206-215 ◽  
Author(s):  
Sylvain Lanouette ◽  
James A. Davey ◽  
Fred Elisma ◽  
Zhibin Ning ◽  
Daniel Figeys ◽  
...  
Keyword(s):  
Protein Design ◽  
Computational Protein Design ◽  
Set Domain ◽  
Lysine Methyltransferase

Electrostatics in computational protein design

2005 ◽  
Vol 9 (6) ◽  
pp. 622-626 ◽  
Author(s):  
Christina L Vizcarra ◽  
Stephen L Mayo
Keyword(s):  
Protein Design ◽  
Computational Protein Design

scholarly journals Protein sequence optimization with a pairwise decomposable penalty for buried unsatisfied hydrogen bonds

2020 ◽  
Author(s):  
Brian Coventry ◽  
David Baker
Keyword(s):  
Aqueous Solution ◽  
Hydrogen Bonds ◽  
Protein Design ◽  
Protein Sequence ◽  
Quadratic Function ◽  
Energetic Cost ◽  
Energy Term ◽  
Sequence Optimization ◽  
Polar Groups ◽  
Sidechain Rotamers

AbstractIn aqueous solution, polar groups make hydrogen bonds with water, and hence burial of such groups in the interior of a protein is unfavorable unless the loss of hydrogen bonds with water is compensated by formation of new ones with other protein groups. Hence, buried “unsatisfied” polar groups making no hydrogen bonds are very rare in proteins. Efficiently representing the energetic cost of unsatisfied hydrogen bonds with a pairwise-decomposable energy term during protein design is challenging since whether or not a group is satisfied depends on all of its neighbors. Here we describe a method for assigning a pairwise-decomposable energy to sidechain rotamers such that following combinatorial sidechain packing, buried unsaturated polar atoms are penalized. The penalty can be any quadratic function of the number of unsatisfied polar groups, and can be computed very rapidly. We show that inclusion of this term in Rosetta sidechain packing calculations substantially reduces the number of buried unsatisfied polar groups.

scholarly journals Structural insights into Escherichia coli phosphopantothenoylcysteine synthetase by native ion mobility–mass spectrometry

2019 ◽  
Vol 476 (21) ◽  
pp. 3125-3139 ◽  
Author(s):  
Daniel Shiu-Hin Chan ◽  
Jeannine Hess ◽  
Elen Shaw ◽  
Christina Spry ◽  
Robert Starley ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Escherichia Coli ◽  
Structural Biology ◽  
Ion Mobility ◽  
Biosynthetic Pathway ◽  
Screening Tool ◽  
Native Mass Spectrometry ◽  
Ion Mobility Mass Spectrometry ◽  
E Coli ◽  
Structural Insights

Abstract CoaBC, part of the vital coenzyme A biosynthetic pathway in bacteria, has recently been validated as a promising antimicrobial target. In this work, we employed native ion mobility–mass spectrometry to gain structural insights into the phosphopantothenoylcysteine synthetase domain of E. coli CoaBC. Moreover, native mass spectrometry was validated as a screening tool to identify novel inhibitors of this enzyme, highlighting the utility and versatility of this technique both for structural biology and for drug discovery.

scholarly journals Identification and biochemical characterization of a novel PP2C-like Ser/Thr phosphatase inE. coli

2018 ◽  
Author(s):  
Krithika Rajagopalan ◽  
Jonathan Dworkin
Keyword(s):  
Biochemical Characterization ◽  
Regulatory Protein ◽  
Biochemical Properties ◽  
Bacterial Genomes ◽  
Phosphatase Inhibitors ◽  
E Coli ◽  
Specificity And Sensitivity ◽  
Genes Encoding ◽  
Number Of Bacteria

AbstractIn bacteria, signaling phosphorylation is thought to occur primarily on His and Asp residues. However, phosphoproteomic surveys in phylogenetically diverse bacteria over the past decade have identified numerous proteins that are phosphorylated on Ser and/or Thr residues. Consistently, genes encoding Ser/Thr kinases are present in many bacterial genomes such asE. coli,which encodes at least three Ser/Thr kinases. Since Ser/Thr phosphorylation is a stable modification, a dedicated phosphatase is necessary to allow reversible regulation. Ser/Thr phosphatases belonging to several conserved families are found in bacteria. One family of particular interest are Ser/Thr phosphatases which have extensive sequence and structural homology to eukaryotic Ser/Thr PP2C phosphatases. These proteins, called eSTPs (eukaryotic-like Ser/Thr phosphatases), have been identified in a number of bacteria, but not inE. coli.Here, we describe a previously unknown eSTP encoded by anE. coliORF,yegK,and characterize its biochemical properties including its kinetics, substrate specificity and sensitivity to known phosphatase inhibitors. We investigate differences in the activity of this protein in closely relatedE. colistrains. Finally, we demonstrate that this eSTP acts to dephosphorylate a novel Ser/Thr kinase which is encoded in the same operon.ImportanceRegulatory protein phosphorylation is a conserved mechanism of signaling in all biological systems. Recent phosphoproteomic analyses of phylogenetically diverse bacteria including the model Gram-negative bacteriumE. colidemonstrate that many proteins are phosphorylated on serine or threonine residues. In contrast to phosphorylation on histidine or aspartate residues, phosphorylation of serine and threonine residues is stable and requires the action of a partner Ser/Thr phosphatase to remove the modification. Although a number of Ser/Thr kinases have been reported inE. coli, no partner Ser/Thrphosphatases have been identified. Here, we biochemically characterize a novel Ser/Thr phosphatase that acts to dephosphorylate a Ser/Thr kinase that is encoded in the same operon.

scholarly journals Rhizobium (Sinorhizobium)meliloti phn Genes: Characterization and Identification of Their Protein Products

1999 ◽  
Vol 181 (2) ◽  
pp. 389-395 ◽  
Author(s):  
George F. Parker ◽  
Timothy P. Higgins ◽  
Timothy Hawkes ◽  
Robert L. Robson
Keyword(s):  
Escherichia Coli ◽  
Sinorhizobium Meliloti ◽  
Insertional Mutagenesis ◽  
Biochemical Characterization ◽  
Polyclonal Antibodies ◽  
E Coli ◽  
New Information ◽  
Phosphorus Sources ◽  
Carbon Phosphorus Lyase

ABSTRACT In Escherichia coli, the phn operon encodes proteins responsible for the uptake and breakdown of phosphonates. The C-P (carbon-phosphorus) lyase enzyme encoded by this operon which catalyzes the cleavage of C-P bonds in phosphonates has been recalcitrant to biochemical characterization. To advance the understanding of this enzyme, we have cloned DNA fromRhizobium (Sinorhizobium) melilotithat contains homologues of the E. coli phnG, -H, -I, -J, and -Kgenes. We demonstrated by insertional mutagenesis that the operon from which this DNA is derived encodes the R. meliloti C-P lyase. Furthermore, the phenotype of this phn mutant shows that the C-P lyase has a broad substrate specificity and that the organism has another enzyme that degrades aminoethylphosphonate. A comparison of the R. meliloti and E. coli phngenes and their predicted products gave new information about C-P lyase. The putative R. meliloti PhnG, PhnH, and PhnK proteins were overexpressed and used to make polyclonal antibodies. Proteins of the correct molecular weight that react with these antibodies are expressed by R. meliloti grown with phosphonates as sole phosphorus sources. This is the first in vivo demonstration of the existence of these hitherto hypothetical Phn proteins.

Sign in / Sign up

Export Citation Format

Share Document