User:Calhobbs11/Protein folding

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Protein Nuclear Magnetic Resonance (NMR) is able to collect protein structural data by inducing a magnet field through samples of concentrated protein. In NMR, depending on the chemical environment, certain nuclei will absorb specific radio-frequencies[1][2]. Because protein structural changes operate on a time scale from ns to ms, NMR is especially equipped to study intermediate structures in timescales of ps to s[3]. Some of the main techniques for studying proteins structure and non-folding protein structural changes include COSY, TOCSY,  HSQC, Time relaxation (T1 & T2), and NOE[1]. NOE is especially useful because as it perturbs hydrogens through magnetization, exchanges between spatially proximal hydrogens are observed[1].

Timescale of protein structural changes matched with NMR experiments. For Protein folding, CPMG Relaxation Dispersion (CPMG RD) and Chemical Exchange Saturation Transfer (CEST) collect data in the appropriate timescale.

Because Protein folding takes place in about 50 to 3000 s−1 CPMG Relaxation dispersion and Chemical exchange Saturation Transfer have become some of the primary techniques for NMR analysis of folding[2]. In addition, both techniques are used to uncover excited intermediate states in the protein folding landscape[4]. To do this, CPMG Relaxation dispersion takes advantage of the Spin echo phenomenon which exposes the target nuclei to a 90 pulse followed by one or more 180 pulses[5]. As the nuclei refocus, a broad distribution indicates the target nuclei is involved in an intermediate excited state. By looking at Relaxation dispersion plots the data collect information on the the thermodynamics and kinetics between the excited and ground[5][4]. Saturation Transfer measures changes in signal from the ground state as excited states become perturbed. It uses weak radio frequency irradiation to saturate the excited state of a particular nuclei which transfers its saturation to the ground state[2]. This signal is amplified by decreasing the magnetization (and the signal) of the ground state[2][4].

The main limitations in NMR is that its resolution decreases sensitivity with proteins that are larger than 25 kDa and in general is not as detailed as X-ray crystallography[2]. Additionally, Protein NMR analysis is quite difficult and can propose multiple solutions from the same NMR spectrum[1].

In a study focused on the folding of an Amyotrophic lateral sclerosis involved protein SOD1, excited intermediates were studied with Relaxation dispersion and Saturation transfer[6]. SOD1 had been previously tied to many disease causing mutants which were supposed to be involved in protein aggregation, however the mechanism was still unknown. By using Relaxation Dispersion and Saturation Transfer experiments many excited intermediate states were uncovered misfolding of SOD1 mutants[6].

  1. ^ a b c d Wüthrich, K. (1990-12-25). "Protein structure determination in solution by NMR spectroscopy". Journal of Biological Chemistry. 265 (36): 22059–22062. ISSN 0021-9258. PMID 2266107.
  2. ^ a b c d e Zhuravleva, Anastasia; Korzhnev, Dmitry M. (2017-05-01). "Protein folding by NMR". Progress in Nuclear Magnetic Resonance Spectroscopy. 100: 52–77. doi:10.1016/j.pnmrs.2016.10.002. ISSN 0079-6565.
  3. ^ Ortega, Gabriel; Pons, Miquel; Millet, Oscar (2013-01-01), Karabencheva-Christova, Tatyana (ed.), "Chapter Six - Protein Functional Dynamics in Multiple Timescales as Studied by NMR Spectroscopy", Advances in Protein Chemistry and Structural Biology, Dynamics of Proteins and Nucleic Acids, vol. 92, Academic Press, pp. 219–251, doi:10.1016/b978-0-12-411636-8.00006-7, retrieved 2020-11-23
  4. ^ a b c Vallurupalli, Pramodh; Bouvignies, Guillaume; Kay, Lewis E. (2012-05-16). "Studying "Invisible" Excited Protein States in Slow Exchange with a Major State Conformation". Journal of the American Chemical Society. 134 (19): 8148–8161. doi:10.1021/ja3001419. ISSN 0002-7863.
  5. ^ a b Neudecker, Philipp; Lundström, Patrik; Kay, Lewis E. (2009-03-18). "Relaxation Dispersion NMR Spectroscopy as a Tool for Detailed Studies of Protein Folding". Biophysical Journal. 96 (6): 2045–2054. doi:10.1016/j.bpj.2008.12.3907. ISSN 0006-3495. PMC 2717354. PMID 19289032.{{cite journal}}: CS1 maint: PMC format (link)
  6. ^ a b Sekhar, Ashok; Rumfeldt, Jessica A. O.; Broom, Helen R.; Doyle, Colleen M.; Sobering, Ryan E.; Meiering, Elizabeth M.; Kay, Lewis E. (2016-11-08). "Probing the free energy landscapes of ALS disease mutants of SOD1 by NMR spectroscopy". Proceedings of the National Academy of Sciences. 113 (45): E6939–E6945. doi:10.1073/pnas.1611418113. ISSN 0027-8424. PMC 5111666. PMID 27791136.{{cite journal}}: CS1 maint: PMC format (link)
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