tag:blogger.com,1999:blog-3300702123878659843.post3436940598086588928..comments2024-03-26T05:25:50.831-04:00Comments on University of Ottawa NMR Facility Blog: HSQC - TOCSYGlenn Faceyhttp://www.blogger.com/profile/05146575170575279335noreply@blogger.comBlogger2125tag:blogger.com,1999:blog-3300702123878659843.post-35249273915672119132016-08-12T12:29:33.347-04:002016-08-12T12:29:33.347-04:00Arifuzzaman,
I'm sorry. I am not an expert in...Arifuzzaman,<br />I'm sorry. I am not an expert in 3D sequences for proteins on Varian instruments. I don' t think I could be of much help. <br />GlennGlenn Faceyhttps://www.blogger.com/profile/05146575170575279335noreply@blogger.comtag:blogger.com,1999:blog-3300702123878659843.post-77370962406888700782016-08-12T11:25:32.855-04:002016-08-12T11:25:32.855-04:00Hello sir,
I need your suggestion about the HSQC...Hello sir, <br /><br />I need your suggestion about the HSQC-TOCSY (or TOCSY-HSQC) experiment (on Varian Inova 600 MHz instrument). <br /><br />As I’m a new user of Varian instrument and also new in protein NMR I'm facing some problems acquiring a 3D TOCSY-HSQC data of a membrane protein. In my current project I need to collect some 3D HSQC-TOCSY (or TOCSY-HSQC) data on Varian Inova 600 MHz instrument for protein side-chain assignment. My sample is a (1.2 m molar) membrane protein inside deuterated DPC micelle in D2O. Its a small membrane protein with MW (ca.) 4 kDa, and considering the micelle the total MW of the protein-micelle system should not exceed 20 kDa.<br /><br /><br />I was trying to use the “gtocsyNhsqc” (with water suppression) pulse sequence to collect the data. However, I couldn’t get any (usable) data using that pulse program, no cross-peak was seen. As the 3D pulse sequence wasn’t working, I ran a 2D ‘gtocsy’ experiment to see the proton-proton correlation but didn’t get any cross-peak also. The 1D data looks good, but in the 2D data, it looks like no polarization transfer is going on. I could not figure out the problem.<br /><br />I tried (for the 2D gtocsy)-<br />>> Mixing time: 30 to 80 m sec<br />>> Relaxation delay: 1 to 2.5 sec<br />>> Spin-locking strength: 6000 (for 10 ppm sweep width)<br /><br />(I can send you the pulse sequence for the 2D gtocsy and the result spectrum)<br /><br /><br />I found the same sample produces nice HSQC 2D spectrum, but for an unknown reason, I’m not getting any (tocsy) cross peak. As I’m new to this instrument and also new in protein NMR, I couldn’t figure out the problem yet. I don't think the problem is with the pulse program, I must doing something wrong in the parameter settings.<br /><br /><br />Could you help me to solve this problem? <b>Any parameter which is important to consider? Any suggestion will be highly appreciated.</b> If needed, I can provide more information.<br /><br />Thanks!!!Anonymoushttps://www.blogger.com/profile/08659457075177103350noreply@blogger.com