I recently was asked a straightforward task: get the energy scores of a given protein by itself, conjugated with fluorescein maleimide and its ring opened form. Despite that I failed miserably because of implicit waters playing odd tricks.
Given that it is of no real use, I am happy to share so to give a tutorial of how to do that in Rosetta (ligand docking), which I found taxing the first time.
Spilt agar on the keyboard
A sticky mess of disconnected ideas, analyses and just plain silly posts.
Monday, 19 March 2018
Saturday, 6 January 2018
Skewed mutational biases: English scrabble with Klingon tiles
Skewed mutational biases: English scrabble with Klingon tiles
In a game of Scrabble one draws seven tiles from a bag and the best results are found when a bag and the language used match. In library making with error-prone PCR one does not have that luxury, but different methods differing degrees of fairness. Mutazyme is like playing in English with a Latin tile set, while manganese Taq with equal nucleotides is like playing in English with a Klingon tile set.Thursday, 4 January 2018
Pi projects: a new year's resolutions
2018 is here. I have papers that I want to or have to write and endless tasks I want to complete, but life is not all about career-driven self-sacrifice, it is also about having fun and what better way than with physical computing. And more time passes the more Rasberry pi projects I find I want to do. So I thought I'd share my shortlist of what I think are the most useful.
Monday, 16 October 2017
Hacking PDBs for fusion protein
This is a How-to guide to make structures of fusion protein —the shoddy way.
NB. This assumes medium–advanced PyMol competency.
NB. This assumes medium–advanced PyMol competency.
Introduction
Representing the structures of fusion protein is a rather annoying task. There are many ways of doing this.
Contrary to expectations, submitting the sequence of a fusion protein to the Phyre server or similar will return spaghetti, just as happens with circular permutations (for which you get a cracked open domain). But give it a go nevertheless —it may save you a lot of time!
Rosetta can be used, but the domains will not be photogenic and it is truly overkill.
So the best way is to simply hack the PDB files and some PyMol ninjutsu.
As an example I will make phusion, i.e. Pyrococcus furiosus DNA polymerase fused to Sulfolobus solfataricus 7D domain (Sso7), because its name basically says it is a fusion.
Contrary to expectations, submitting the sequence of a fusion protein to the Phyre server or similar will return spaghetti, just as happens with circular permutations (for which you get a cracked open domain). But give it a go nevertheless —it may save you a lot of time!
Rosetta can be used, but the domains will not be photogenic and it is truly overkill.
So the best way is to simply hack the PDB files and some PyMol ninjutsu.
As an example I will make phusion, i.e. Pyrococcus furiosus DNA polymerase fused to Sulfolobus solfataricus 7D domain (Sso7), because its name basically says it is a fusion.
Friday, 18 August 2017
Rosetta easteregg
The guide to use Rosetta may be a bit, ehm, flaky, but I think it is made up for by this rather amusing comment header I found in one of its
Perl script, kudos to the author.
############################################################################### # # MAKE_FRAGMENTS.PL 1.00 -- THE (PEN)ULTIMATE IN HOME FRAGMENT-PICKING SOFTWARE! # # CAUTION: NO USER SERVICEABLE PARTS BELOW! # # TO REDUCE RISK OF ELECTRIC SHOCK, DO NOT REMOVE THE COVER! # DO NOT ATTEMPT REPAIRS! REFER SERVICING TO YOUR AUTHORIZED DEALER! # AVOID PROLONGED EXPOSURE TO HEAT OR SUNLIGHT! # TO REDUCE THE RISK OF FIRE OR ELECTRIC SHOCK, DO NOT EXPOSE THE # PRODUCT TO RAIN AND/OR MOISTURE! # DO NOT MOVE THE PRODUCT WHILE IN USE! # DO NOT LOOK AT THE PRODUCT WHILE IN USE! # DO NOT COMPLAIN ABOUT THE PRODUCT WHILE IN USE! # DO NOT DISCUSS THE PRODUCT WHILE IN USE! # DO NOT THINK ABOUT THE PRODUCT WHILE IN USE! # CLEAN ONLY WITH MILD DETERGENTS AND A SOFT CLOTH! # USE ONLY IN WELL-VENTILATED AREAS! # # FOR EXTERNAL USE ONLY! DO NOT TAKE INTERNALLY! # MAY PRODUCE STRONG MAGNETIC FIELDS! # # DO NOT REMOVE THIS TAG UNDER PENALTY OF LAW. # # THIS ARTICLE CONTAINS NEW MATERIAL ONLY. # # THIS LABEL IS AFFIXED IN COMPLAINCE WITH THE UPHOLSTERED AND # STUFFED ARTICLES ACT. # # # (IN OTHER WORDS: DON'T EVEN *THINK* ABOUT CHANGING THINGS BELOW THIS POINT!) # ###############################################################################
Wednesday, 5 July 2017
In vivo shuffling via heteroduplex amplicons
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Bluescreen transilluminator photo showing different shades of green. |
I did a sneaky experiment to see if this could be used to make protocol to shuffle mutations between variants and got a positive result, but possibly not as effective as hoped.
Sunday, 23 April 2017
A note on Mutazyme and Manganese
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