I would like to start the “journal club” section of the NMR forum by presenting my favorite paper from the recent literature.
The paper (in this weeks issue of Science) is, in my opinion at least, a perfect example of the ability of NMR to contribute to the structural analysis of a challenging RNA-protein complex
I chose this high-impact paper because the combination of biochemistry, NMR, and crystallography on a ternary complex is how I would like to tackle a challenging project that I am working on.
The authors used chemical shift perturbation and saturation transfer to monitor the effect of titrating Prp31 into the preformed U4-15.5K RNA-protein complex. To do this the authors purified nitrogen-15 labeled and deuterated samples of the 15.5K construct. All of the proteins were produced in E.coli using well-established protocols. Gel filtration was used to purify the complexes to yield homogeneous samples
Subsequent crystallogrpahic analysis confirmed formation of the ternary complex and provides a rationale for how Prp31 contributes to the stability of the complex. The structural studies in combination with biochemical experiments yield valuable insights into spliceosome mechanism. NMR experiments such as the ones reported here are well within the reach of most laboratories with access to instrumentation and can be very powerful in lending confidence to subsequent crystallization experiments.
[NMR paper] Rotational-echo double-resonance NMR-restrained model of the ternary complex of 5-eno
Rotational-echo double-resonance NMR-restrained model of the ternary complex of 5-enolpyruvylshikimate-3-phosphate synthase.
Related Articles Rotational-echo double-resonance NMR-restrained model of the ternary complex of 5-enolpyruvylshikimate-3-phosphate synthase.
J Biomol NMR. 2004 Jan;28(1):11-29
Authors: McDowell LM, Poliks B, Studelska DR, O'Connor RD, Beusen DD, Schaefer J
The 46-kD enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase catalyzes the condensation of shikimate-3-phosphate (S3P) and phosphoenolpyruvate to form EPSP....
nmrlearner
Journal club
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11-24-2010 09:25 PM
[NMR paper] NMR structure of the 3' stem-loop from human U4 snRNA.
NMR structure of the 3' stem-loop from human U4 snRNA.
Related Articles NMR structure of the 3' stem-loop from human U4 snRNA.
Nucleic Acids Res. 2002 Oct 15;30(20):4371-9
Authors: Comolli LR, Ulyanov NB, Soto AM, Marky LA, James TL, Gmeiner WH
The NMR structure of the 3' stem-loop (3'SL) from human U4 snRNA was determined to gain insight into the structural basis for conservation of this stem-loop sequence from vertebrates. 3'SL sequences from human, rat, mouse and chicken U4 snRNA each consist of a 7 bp stem capped by a UACG tetraloop. No...
nmrlearner
Journal club
0
11-24-2010 08:58 PM
[NMR paper] NMR solution structure of the antitumor compound PT523 and NADPH in the ternary compl
NMR solution structure of the antitumor compound PT523 and NADPH in the ternary complex with human dihydrofolate reductase.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--pubs.acs.org-images-acspubs.jpg Related Articles NMR solution structure of the antitumor compound PT523 and NADPH in the ternary complex with human dihydrofolate reductase.
Biochemistry. 1997 Apr 15;36(15):4399-411
Authors: Johnson JM, Meiering EM, Wright JE, Pardo J, Rosowsky A, Wagner G
The antitumor compound PT523 was found to have an inhibition constant (K(i))...
nmrlearner
Journal club
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08-22-2010 03:31 PM
[NMR paper] NMR solution structure of the antitumor compound PT523 and NADPH in the ternary compl
NMR solution structure of the antitumor compound PT523 and NADPH in the ternary complex with human dihydrofolate reductase.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--pubs.acs.org-images-acspubs.jpg Related Articles NMR solution structure of the antitumor compound PT523 and NADPH in the ternary complex with human dihydrofolate reductase.
Biochemistry. 1997 Apr 15;36(15):4399-411
Authors: Johnson JM, Meiering EM, Wright JE, Pardo J, Rosowsky A, Wagner G
The antitumor compound PT523 was found to have an inhibition constant (K(i))...
nmrlearner
Journal club
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08-22-2010 03:03 PM
[NMR paper] Evidence for a ternary complex formed between flavodoxin and cytochrome c3: 1H-NMR an
Evidence for a ternary complex formed between flavodoxin and cytochrome c3: 1H-NMR and molecular modeling studies.
Related Articles Evidence for a ternary complex formed between flavodoxin and cytochrome c3: 1H-NMR and molecular modeling studies.
Biochemistry. 1994 May 31;33(21):6394-407
Authors: Palma PN, Moura I, LeGall J, Van Beeumen J, Wampler JE, Moura JJ
Small electron-transfer proteins such as flavodoxin (16 kDa) and the tetraheme cytochrome c3 (13 kDa) have been used to mimic, in vitro, part of the complex electron-transfer chain...
nmrlearner
Journal club
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08-22-2010 03:33 AM
[NMR paper] Evidence for a ternary complex formed between flavodoxin and cytochrome c3: 1H-NMR an
Evidence for a ternary complex formed between flavodoxin and cytochrome c3: 1H-NMR and molecular modeling studies.
Related Articles Evidence for a ternary complex formed between flavodoxin and cytochrome c3: 1H-NMR and molecular modeling studies.
Biochemistry. 1994 May 31;33(21):6394-407
Authors: Palma PN, Moura I, LeGall J, Van Beeumen J, Wampler JE, Moura JJ
Small electron-transfer proteins such as flavodoxin (16 kDa) and the tetraheme cytochrome c3 (13 kDa) have been used to mimic, in vitro, part of the complex electron-transfer chain...
nmrlearner
Journal club
0
08-22-2010 03:33 AM
[Nature network NMR forum] Journal club: structural biology of HIV infection (0 replies)
Journal club: structural biology of HIV infection (0 replies)
A recent paper in Science highlights once again the potential of NMR to yield insights into complex molecular recognition events in critical biological processes.
In this case the authors used a combination of X-ray crystallography and NMR to probe the structure of the extracellular terminus of the CCR5 co-receptor which, in concert with human CD4 and HIV gp120, is crucial for viral entry. The authors applied challenging saturation transfer effects to study the structure of a small synthetic peptide in a ternary...
nmrlearner
News from other NMR forums
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08-21-2010 03:29 PM
[Nature network NMR forum] Journal club: recombinant RNA expression (0 replies)
Journal club: recombinant RNA expression (0 replies)
Recently I was thinking about my RNA project and wishing that I could express RNA in bacteria as is typically done for recombinant proteins. I looked at the July issue of Nature Methods to see a paper describing exactly the technology I was imagining. The authors created plasmids which allow the fusion of a target RNA sequence to a tRNA scaffold which is authenically processed by the cellular machinery and protected from degradation by the intrinsic stability of the tRNA. The potential benefits of this technology for NMR of RNA...