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Enzyme Design

fastdesign with constraints in RosettaScripts

Category: 
Enzyme Design

Hi,

I am using the FastDesign protocol in RosettaScripts to design some residues in the neighborhood of a ligand, specifically with constraints of N-O pair distance for NAD binding. 

I've added constraints for the two N-O pairs to be 3.0 and 3.3 angstroms each with harmonic function and sd of 0.5, but somehow the distances are not kept below ~4.0 at all. (I've just observed a structure with over 6.0 angstrom distances for the two pairs.

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Error when running enzyme_design tutorial

Category: 
Enzyme Design

I'm trying to run the enzyme_design tutorial from here:

 

https://www.rosettacommons.org/demos/latest/public/enzyme_design/README

The first 2 stages run fine, but when I run the Design step with the command:

$ROSETTA3/bin/enzyme_design.static.linuxgccrelease @rosetta_inputs/general_design.flags -s rosetta_inputs/UM_1_D41H116K189_1tml_11_mocktim_1.pdb -out:file:o scorefile.txt

The code crashed with the following error:

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How tu use Delta filter's relax_mover

Category: 
Enzyme Design

Hi,

I want to use https://www.rosettacommons.org/docs/latest/scripting_documentation/RosettaScripts/Filters/filter_pages/DeltaFilter to calculate the difference in energy between wt and design. I think I have to use parameter relax_mover, but from the docs It is not clear what value to use. Does anyone have any experience with it?

Edit:

I dug into the source code and it appears that this should work:

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Re: Glutamic acid protonation

Category: 
Enzyme Design

Hi,

I am working on an enzyme design project for which I want one of the glutamic acid of the catalytic site to be protonated at OE2 position. I manually did that using schrodinger but when I prepared the structure using relax protocol, the glutamic acid becomes deprotonated. How, I can get a specific glutamic acid (For eg position 43) to be protonated specifically at OE2?

 

Thanks in advance for help

BH

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Altering substrate specificity

Category: 
Design
Scoring
Enzyme Design
Small Molecules
Chemically Modified Residues

Hi all,

I'm presently working on a project to modify the substrate specificity of a DNA polymerase for non-natural nucleic acids.

I've currently been using the GreedyOptMutationMover with a ddg filter (jump across the substrate) for a 10Å shell around the active site and identified several single point mutations that have been experimentally validated to improve activity.

However, one of the problems with greedyopt is that it does not appear to screen every possible position or single point mutation within the designable region.

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RosettaMatch outputs

Category: 
Design
Enzyme Design

I've been running several rounds of RosettaMatch lately and I'd like them to go faster. I'm pretty sure writing matches is the part slowing it down. Is there anyway to limit the number of matches it outputs? For example, for the same matched sequence (say H133H135H432), I get anywhere from 50-400 cloudPDB files. I don't want more than the first one or two, because the rest are almost always just different ligand positions and I'll end up running EnzDesign anyways. I'm using these flags:

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Enzyme Design with additional covalent backbone bonds

Category: 
Design
Enzyme Design
Constraints
Non-Canonical Peptides

Hey everyone,

I am trying to design a protein with a covalent bond between the C of Thr and the N of Gly with a Tyr in between them (imitating the chromophore from GFP for perspective). Because of this odd 5-membered ring formation, I used Thr/Gly with a generous distance constraint between them in my CST file when I ran RosettaMatch to find likely sites in my scaffold proteins. My sites place the two backbone atoms ~4-5A apart.

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