BioNMR
NMR aggregator & online community since 2003
BioNMR    
Learn or help to learn NMR - get free NMR books!
 

Go Back   BioNMR > Educational resources > Journal club
Advanced Search



Jobs Groups Conferences Literature Pulse sequences Software forums Programs Sample preps Web resources BioNMR issues


Webservers
NMR processing:
MDD
NMR assignment:
Backbone:
Autoassign
MARS
UNIO Match
PINE
Side-chains:
UNIO ATNOS-Ascan
NOEs:
UNIO ATNOS-Candid
UNIO Candid
ASDP
Structure from NMR restraints:
Ab initio:
GeNMR
Cyana
XPLOR-NIH
ASDP
UNIO ATNOS-Candid
UNIO Candid
Fragment-based:
BMRB CS-Rosetta
Rosetta-NMR (Robetta)
Template-based:
GeNMR
I-TASSER
Refinement:
Amber
Structure from chemical shifts:
Fragment-based:
WeNMR CS-Rosetta
BMRB CS-Rosetta
Homology-based:
CS23D
Simshift
Torsion angles from chemical shifts:
Preditor
TALOS
Promega- Proline
Secondary structure from chemical shifts:
CSI (via RCI server)
TALOS
MICS caps, β-turns
d2D
PECAN
Flexibility from chemical shifts:
RCI
Interactions from chemical shifts:
HADDOCK
Chemical shifts re-referencing:
Shiftcor
UNIO Shiftinspector
LACS
CheckShift
RefDB
NMR model quality:
NOEs, other restraints:
PROSESS
PSVS
RPF scores
iCing
Chemical shifts:
PROSESS
CheShift2
Vasco
iCing
RDCs:
DC
Anisofit
Pseudocontact shifts:
Anisofit
Protein geomtery:
Resolution-by-Proxy
PROSESS
What-If
iCing
PSVS
MolProbity
SAVES2 or SAVES4
Vadar
Prosa
ProQ
MetaMQAPII
PSQS
Eval123D
STAN
Ramachandran Plot
Rampage
ERRAT
Verify_3D
Harmony
Quality Control Check
NMR spectrum prediction:
FANDAS
MestReS
V-NMR
Flexibility from structure:
Backbone S2
Methyl S2
B-factor
Molecular dynamics:
Gromacs
Amber
Antechamber
Chemical shifts prediction:
From structure:
Shiftx2
Sparta+
Camshift
CH3shift- Methyl
ArShift- Aromatic
ShiftS
Proshift
PPM
CheShift-2- Cα
From sequence:
Shifty
Camcoil
Poulsen_rc_CS
Disordered proteins:
MAXOCC
Format conversion & validation:
CCPN
From NMR-STAR 3.1
Validate NMR-STAR 3.1
NMR sample preparation:
Protein disorder:
DisMeta
Protein solubility:
camLILA
ccSOL
Camfold
camGroEL
Zyggregator
Isotope labeling:
UPLABEL
Solid-state NMR:
sedNMR


Reply
Thread Tools Search this Thread Rate Thread Display Modes
  #1  
Unread 03-24-2019, 10:41 PM
nmrlearner's Avatar
Senior Member
 
Join Date: Jan 2005
Posts: 23,137
Points: 193,617, Level: 100
Points: 193,617, Level: 100 Points: 193,617, Level: 100 Points: 193,617, Level: 100
Level up: 0%, 0 Points needed
Level up: 0% Level up: 0% Level up: 0%
Activity: 50.7%
Activity: 50.7% Activity: 50.7% Activity: 50.7%
Last Achievements
Award-Showcase
NMR Credits: 0
NMR Points: 193,617
Downloads: 0
Uploads: 0
Default Alpha protons as NMR probes in deuterated proteins

Alpha protons as NMR probes in deuterated proteins

Abstract

We describe a new labeling method that allows for full protonation at the backbone Hα position, maintaining protein side chains with a high level of deuteration. We refer to the method as alpha proton exchange by transamination (α-PET) since it relies on transaminase activity demonstrated here using Escherichia coli expression. We show that α-PET labeling is particularly useful in improving structural characterization of solid proteins by introduction of an additional proton reporter, while eliminating many strong dipolar couplings. The approach benefits from the high sensitivity associated with 1.3Â*mm samples, more abundant information including Hα resonances, and the narrow proton linewidths encountered for highly deuterated proteins. The labeling strategy solves amide proton exchange problems commonly encountered for membrane proteins when using perdeuteration and backexchange protocols, allowing access to alpha and all amide protons including those in exchange-protected regions. The incorporation of Hα protons provides new insights, as the close Hαâ??Hα and Hαâ??HN contacts present in β-sheets become accessible, improving the chance to determine the protein structure as compared with HNâ??HN contacts alone. Protonation of the Hα position higher than 90% is achieved for Ile, Leu, Phe, Tyr, Met, Val, Ala, Gln, Asn, Thr, Ser, Glu, Asp even though LAAO is only active at this degree for Ile, Leu, Phe, Tyr, Trp, Met. Additionally, the glycine methylene carbon is labeled preferentially with a single deuteron, allowing stereospecific assignment of glycine alpha protons. In solution, we show that the high deuteration level dramatically reduces R2 relaxation rates, which is beneficial for the study of large proteins and protein dynamics. We demonstrate the method using two model systems, as well as a 32Â*kDa membrane protein, hVDAC1, showing the applicability of the method to study membrane proteins.



Source: Journal of Biomolecular NMR
Reply With Quote


Did you find this post helpful? Yes | No

Reply
Similar Threads
Thread Thread Starter Forum Replies Last Post
Electron spin resonance studies on deuterated nitroxyl spin probes used in Overhauser-enhanced magnetic resonance imaging #DNPNMR #ODNP
From The DNP-NMR Blog: Electron spin resonance studies on deuterated nitroxyl spin probes used in Overhauser-enhanced magnetic resonance imaging #DNPNMR #ODNP Jebaraj, D. David, Hideo Utsumi, and A. Milton Franklin Benial. “Electron Spin Resonance Studies on Deuterated Nitroxyl Spin Probes Used in Overhauser-Enhanced Magnetic Resonance Imaging.” Magnetic Resonance in Chemistry 55, no. 8 (2017): 700–705. https://doi.org/10.1002/mrc.4576.
nmrlearner News from NMR blogs 0 06-23-2018 12:47 AM
Dynamic nuclear polarization studies on deuterated nitroxyl spin probes #DNPNMR
From The DNP-NMR Blog: Dynamic nuclear polarization studies on deuterated nitroxyl spin probes #DNPNMR p.p1 {margin: 0.0px 0.0px 0.0px 36.0px; text-indent: -36.0px; font: 12.0px Helvetica} David Jebaraj, D., H. Utsumi, and A. Milton Franklin Benial, Dynamic nuclear polarization studies on deuterated nitroxyl spin probes. Magn Reson Chem, 2017. 55(10): p. 909-916. https://www.ncbi.nlm.nih.gov/pubmed/28444914
nmrlearner News from NMR blogs 0 10-03-2017 03:24 AM
[NMR paper] Access to aliphatic protons as reporters in non-deuterated proteins by solid-state NMR.
Access to aliphatic protons as reporters in non-deuterated proteins by solid-state NMR. http://www.bionmr.com//www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www.rsc.org-images-entities-char_z_RSClogo.gif Related Articles Access to aliphatic protons as reporters in non-deuterated proteins by solid-state NMR. Phys Chem Chem Phys. 2015 Dec 21; Authors: Vasa SK, Rovó P, Giller K, Becker S, Linser R Abstract Interactions within proteins, with their surrounding, and with other molecules are mediated mostly by hydrogen atoms. In...
nmrlearner Journal club 0 12-28-2015 12:26 AM
An optimized method for 15 N R 1 relaxation rate measurements in non-deuterated proteins
An optimized method for 15 N R 1 relaxation rate measurements in non-deuterated proteins Abstract 15N longitudinal relaxation rates are extensively used for the characterization of protein dynamics; however, their accurate measurement is hindered by systematic errors. 15N CSA/1Hâ??15N dipolar cross-correlated relaxation (CC) and amide proton exchange saturation transfer from water protons are the two main sources of systematic errors in the determination of 15N R1 rates through 1Hâ??15N HSQC-based experiments. CC is usually...
nmrlearner Journal club 0 05-07-2015 12:59 AM
[NMR paper] Assignment of the nonexchanging protons of the alpha-spectrin SH3 domain by two- and
Assignment of the nonexchanging protons of the alpha-spectrin SH3 domain by two- and three-dimensional 1H-13C solid-state magic-angle spinning NMR and comparison of solution and solid-state proton chemical shifts. Related Articles Assignment of the nonexchanging protons of the alpha-spectrin SH3 domain by two- and three-dimensional 1H-13C solid-state magic-angle spinning NMR and comparison of solution and solid-state proton chemical shifts. Chembiochem. 2001 Dec 3;2(12):906-14 Authors: van Rossum BJ, Castellani F, Rehbein K, Pauli J, Oschkinat H ...
nmrlearner Journal club 0 11-19-2010 08:44 PM
[NMR paper] NMR spectroscopy of hydroxyl protons in aqueous solutions of peptides and proteins.
NMR spectroscopy of hydroxyl protons in aqueous solutions of peptides and proteins. Related Articles NMR spectroscopy of hydroxyl protons in aqueous solutions of peptides and proteins. J Biomol NMR. 1992 Sep;2(5):447-65 Authors: Liepinsh E, Otting G, Wüthrich K Hydroxyl groups of serine and threonine, and to some extent also tyrosine are usually located on or near the surface of proteins. NMR observations of the hydroxyl protons is therefore of interest to support investigations of the protein surface in solution, and knowledge of the...
nmrlearner Journal club 0 08-21-2010 11:45 PM
NMR of Naphthalene: why are the alpha-protons more downfield than the beta- protons?
Hi, can you please help me explain why the alpha-protons of naphthalene are further downfield? I know that the protons at the alpha position must be more deshielded, but I don't know how to explain why they have less electron density compared to the beta protons. Does this have to do with the number of double bonds that can be drawn in different resonance structures? Thanks for your help!Thanks so much!
flan318 NMR Questions and Answers 1 12-24-2002 12:07 PM


Thread Tools Search this Thread
Search this Thread:

Advanced Search
Display Modes Rate This Thread
Rate This Thread:

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is On
Trackbacks are Off
Pingbacks are Off
Refbacks are Off



BioNMR advertisements to pay for website hosting and domain registration. Nobody does it for us.



Powered by vBulletin® Version 3.7.3
Copyright ©2000 - 2024, Jelsoft Enterprises Ltd.
Copyright, BioNMR.com, 2003-2013
Search Engine Friendly URLs by vBSEO 3.6.0

All times are GMT. The time now is 03:39 PM.


Map