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by: Adah McCullough III


Adah McCullough III
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This 8 page Class Notes was uploaded by Adah McCullough III on Saturday September 12, 2015. The Class Notes belongs to BCMB 4190 at University of Georgia taught by Staff in Fall. Since its upload, it has received 16 views. For similar materials see /class/202193/bcmb-4190-university-of-georgia in Biochemistry and Molecular Biology at University of Georgia.

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Date Created: 09/12/15
CHEM BCMB 419061908189 Introductory NMR Lecture 19 CHEM 41906190 The NOESY Experiment 1 Review ID steadystate NOE experiment 90 x I t1 used in cases of well resolved spectra for assignment purposes or to establish local or global geometry by saturation of selected signals the steadystate NOE develops during saturation of one spin and results in a change in the intensity of the signal from another spin this is in contrast to the transient NOE see below X A saturate pm X E A used most often these days for enhancement of 13C signals by broadband decoupling of 1H FID 13C 2 1D and 2D transient NOE experiments NOESY a 1D selective transient NOE experiment an NOE can also be induced by application of a selective 180 pulse to one spin the A spin 180 x 90 x t1 during the mixing time cm the NOE starts to build at the X spin as the W0 and W2 routes redistribute the spin populations the NOE is transient however it builds to a maximum in a short time approximately one T1 period and then decreases remember only a few T1 periods after a 180 pulse equilibrium populations are established b 2D NOE experiment NOESY preparation evolution mixing detection t1 t2 90 90 COSY t1 H 90 90 90 NOESY t1 Em 90 TOCSY t1 90x 180x 90y 180 90 180 H Ht Wm H r r i HSQC 180x 90x E 90 180 13C t12 i t12 39 decouple 3 the NOESY Euclear Qverhauser Effect Spectroscopy experiment plays a central role in the structural elucidation of molecules ranging from small organic molecules to large biological molecules nucleic acids and proteins 90 x 90 x 90 x t2 in the 2D NOESY experiment all spins are inverted by the first two 90 pulses assume for the moment that t1 0 permitting all transient NOE s to develop during the subsequent mixing time 17m increasing t1 provides chemical shift modulation of the signal for the second dimension in the 2D NOESY spectrum the peaks representing the transferred magnetization the NOEs appear as crosspeaks offdiagonal peaks the crosspeaks appear at the chemical shifts of the spins involved the intensities of the crosspeaks are much less than the intensities of the diagonal peaks 3 2D ESY vector dia rams ewe again will assume a simple AX system with no coupling JAX 0 A X A Z 6 W V x Z f y y X A and X have different Larmor frequencies so M A and MX precess about z and move apart during t1 the second 900 pulse converts the y components ofMA and MX to z during 1 polarization is transferred from A to X and from X to A via crossrelaxation W0 and W2 the transferred components are represented by the small arrows with opposite coloring 4 2D NOESY spectrum I o 39 X F1 lt1Hppmgt o A f l I A X F2 1H ppm Crossrelaxation crosspolarization between nuclei results in cross peaks in the 2D NOESY spectrum usually referred to as NOE crosspeaks or simply NOEs the intensity of the cross peaks is substantially less than the intensity of the diagonal peaks the diagonal peaks can be problematic in these eXperiments because they are very intense the intensity of the cross peaks is proportional to 1r6 Where r is the internuclear distance Ieclr6 so for 1H H distances if the distance is 5 approximately 5 A crosspeaks can be observed the sign or phase of crosspeaks relative to the diagonal peaks depends on the size of the molecule for small molecules the crosspeaks and diagonal peaks are of opposite phase for large molecules the crosspeaks are the same phase as the diagonal peaks 5 2D ESY role of l and Q gradient pulses Normally the data from many scans at each t1 are added together during collection of N OESY spectra ideally one would wait 5T1 between scans to allow the system to equilibrate this of course require far too much time to acquire a complete 2D data set one can wait for lt5T1 between scans but this causes a loss in signal intensity and an increase in the appearance of artifacts in the 2D spectra some of which can be alleviated by phase cycling and other techniques linear eld gradient pulses can be used to G1 gradient in the gradient N OESY experiment the G1 gradient pulse will dephase any transverse magnetization during the recycle delay preventing artifacts in the spectra resulting from these components Gz gradient performs a function similar to the G1 gradient only during the mixing time 1 During 1m transverse magnetization components often can cause artifacts in NOESY spectra G2 dephases these components 2D ESY mixin time How long should the mixing time 1 be 1quot5 in order to maximize the polarization 01 TI transfer that can be accomplished by the 1039 transient NOE and therefore to maximize the crosspeak intensity in the NOESY I spectrum 1quotI should be set to nI approximately T1 for the 1H spins involved 04 T2 small molecule 500 ms 5 s 101 large molecule 30 ms 400 ms 10 10quot 10quot 1039 1oquotl Tcs 7 exam le 1D stead state Eex eriment 3 CH Which methyl signal 517 or 515 ppm is the methyl group on the same side of the ring as H3 saturate the methyl signals when the signal at 517 ppm is saturated an NOE enhancement is observed for H3 indicating the signal at 517 ppm corresponds to the methyl group on the same side of the ring as H3


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