My software notes

GUARDD: user-friendly MATLAB software for rigorous analysis of CPMG RD NMR data

kpwu — Tue, 20 Dec 2011 22:49:38 +0000

There is an article published at Journal of Biomolecular NMR recently which describes a package of MATLAB scripts to analyze CPMG relaxation dispersion NMR data. I haven’t done CPMG RD experiments on my target proteins, however, I know the analysis of CPMG RD data is not trivial. This program, GUARDD, seems to be a very useful and UI-friendly toolkit to efficiently obtain information of Rex, Kex and other relaxation rates from CPMG RD data.

The following snapshots are directly linked to GUARDD homepage.

More information about GUARDD:

Title: : GUARDD: user-friendly MATLAB software for rigorous analysis of CPMG RD NMR data.
Journal: Journal of Biomolecular NMR (online published first, Dec.20. 2011)
Homepage at google code: http://code.google.com/p/guardd/

Additional information:
Matlab, a product from Mathworks
CATIA (Cpmg, Anti-trosy, and Trosy Intelligent Analysis) from Lewis Kay’s group
CPMGFit from Arther Palmer’s group

[PyMOL] gradient background color

kpwu — Tue, 15 Nov 2011 17:03:56 +0000

Starting at PyMOL 1.4, single color for the background is no longer the only choice. By using 3 PyMOL commands, the gradient background color can be easily shown. The only thing users have to know is that command “ray” doesn’t support this gradient color yet. So if you type “ray” in the pymol terminal , the gradient color will show off.

Here is what I did (use PyMOL 1.4) at get the following picture:

set bg_rgb_top, [0,255,0]
set bg_rgb_bottom, [0,255,255]
set bg_gradient, on
png gradient_bg.png

[PyMOL] Sidechain helper in the cartoon view

kpwu — Sat, 12 Nov 2011 20:25:27 +0000

Using cartoon view to present the protein secondary structures and tertiary structure is common and we also often see people present some sidechains of particular residues which may be important in some biological roles. In PyMOL, when sidechain atoms are presented in the “sticks mode” with the cartoon scaffold, we always see the backbone atoms (e.g. N, O) are also presented in the sticks mode.

Here is the example (PDB code: 2BTF) showing that the nitrogen (blue) of Val47 is displaying inside the helix.

By using a PyMOL commend “set cartoon_side_chain_helper”, we can quickly hide sticks of the nitrogen and oxygen atoms.

Command: set catoon_side_chain_helper, on

note: “on” for removing the sticks, “off” (default argument) is to keep all atoms in sticks view

Note (11/14/2011): PyMOLWiki has an example page of this command, see here.

NMR protocol (recipe)–make stretch gel for protein RDC measurement

kpwu — Wed, 19 Oct 2011 23:01:03 +0000

I bought the starter kit (NE-373-B-6/4.2) at NewEraNMR for RDC experiment on Varian NMR (with 700 MHz grade) and I decided to stretch the gel a lot, so 6mm-wide gel chamber and “6mm-4.2mm gel funnel” are included.

The NMR tube seems to be more fragile than Wilmad products I often used, so I also ordered additional 5 tubes (both ends are opened).

Since the special point of the NMR tubes used for this stretch gel is “two-end opened”, one can find glass factory to cut the regular NMR then no need to buy this special tube.

To users who can use both Varian and Bruker NMR machines: you just need to buy some “End Gel Plug” for Bruker machine. It’s not necessary to buy two starter kits.

The picture shown left is an example I made and the bubble near the bottom of gel showed up after 3 days. User may have to to worry the time-dependent changes of the stretched gel.

Required solution:

30% acrylamide (w/v) in distilled H2O
0.8% bis-acrylamide (w/v) in distilled H2O
10% APS (w/v), APS = ammonium persulfate
TEMED as catalyst
Commercial kit to make stretch gel in NMR tube (page 16 in NewEraNMR’s catalog)

Recipe for 1ml mixture: (ul used at here means micro-liter)

6% gel:

30% acrylamide: 200 ul
1% bis-acrylamide: 80 ul
10% APS: 10 ul
TEMED: 0.1 ul
H2O: 710

4.2 % gel:

30% acrylamide: 140 ul
1% bis-acrylamide: 56 ul
10% APS: 8 ul
TEMED: 0.1 ul
H2O: 796

You may note that I made the ratios of acrylamide:bis-acrylamide:APS constant between 6% and 4.2% gel. Just try to avoid too much “unreacted” APS and acrylamide inside the gel.

Protocol of gel preparation:

Mix the above recipe and load 800 ul into the gel chamber, wait for 2 hour for gel formation. The “un-used” 200 ul mixture is used to check if the gel is formed.
Gently remove the gel from chamber using the extrude rod provided in the starter kit and use cleaned knife to cut the gel to a proper length. I made both 1.5 and 2mm gels to next step.
Wash the short gels with 15 ml distilled water for 1 hour and repeat 3-5 times. This step is to try to remove unreacted acrylamide, TEMED, and APS inside the gel.
Wash the gel with 15 ml NMR experimental buffer for 1 hour, repeat 3 times. (or do more volume/times and skip step 3)
Soak the gel with 600 ul NMR buffer which has 10% D2O inside and leave it overnight.
Remove the buffer and transfer~300 ul concentrated NMR sample (10% D2O included) to the tube containing short gel.Wait for free diffusion for overnight.
I usually use ~1 mM NMR sample and the final protein concentration left in the solution is less than 600 uM suggesting 300-400 uM protein are inside the gel.
Assemble the gel chamber with funnel, load the short gel (with protein and 10% D2O) into it. Follow the guide in the NewEra’s catalogue, it should be easy to move the gel from chamber to NMR tube.
Once the gel is fully inserted into the NMR tube, it’s stretched and quickly insert the “End Gel Plug” then insert the assembled “rod-end up plug” to keep the gel at proper position.

As my personal experiences, 6% gel and 4.2% gel returned me 2H splitting constants of 6.2 Hz and 2.5Hz, respectively.

Bruker’s Topspin running on Mac

kpwu — Wed, 19 Oct 2011 18:21:22 +0000

Balance the report as I just made a post for Agilent’s VnmrJ.

I found from a web blog article that introduces the news–Bruker’s Topspin 3.1 for Mac.

Bruker’s brief introduction of Topspin for Mac machine includes features:

Mac OS X platform for TopSpin processing
Inspire a new generation of NMR users, experts and students alike
Entirely programmed in the native Apple Mac OS X environmentData analysis, processing and simulation features for efficient small molecule and structural biology research
Ideal data processing and analysis aid for NMR training

Snapshots of Topspin running on Mac linked to Bruker’s example images show at here:

According to Bruker’s purchase link, the price is $99 of academic license for one Mac machine. The computer MAC address code must be provided meaning one license is only available for one Mac machine!!

The current version (Oct of 2011) Topspin for Mac required “Core 2 Duo” type CPU and OS 10.6 (snow leopard and later). My old macbook pro with “core duo” is not available this time. Sad. I can only try to install it on my iMac.

Maybe I should email Bruker to request something like 30-60 days free trial and make some blog articles for “Bruker Topspin on Mac” as an exchange.