Large Protein Complexes with Extreme Rotational Correlation Times Investigated in Solution by Magic-Angle-Spinning NMR Spectroscopy

Mainz, Andi; Jehle, Stefan; Rossum, Barth van; Oschkinat, Hartmut; Reif, Bernd

doi:10.1021/ja904733v

Cited by 89 publications

(131 citation statements)

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“…Solidstate NMR provides atomic resolution structural information of frozen protein solutions. Therefore, solid-state NMR has been applied to study protein folding (26) and large protein complexes (27) in frozen solution, as well as AFPs bound to ice (28)(29)(30). Recently, we confirmed the location of the ice-binding surface of AFP III by comparing liquid-state NMR spectra of this protein in solution with solid-state NMR 2D spectra of AFP III in frozen solution (6).…”

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confidence: 64%

Protein–ice interaction of an antifreeze protein observed with solid-state NMR

Siemer

Huang

McDermott

2010

Proc. Natl. Acad. Sci. U.S.A.

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NMR on frozen solutions is an ideal method to study fundamental questions of macromolecular hydration, because the hydration shell of many biomolecules does not freeze together with bulk solvent. In the present study, we present previously undescribed NMR methods to study the interactions of proteins with their hydration shell and the ice lattice in frozen solution. We applied these methods to compare solvent interaction of an ice-binding type III antifreeze protein (AFP III) and ubiquitin a non-ice-binding protein in frozen solution. We measured 1 H-1 H cross-saturation and cross-relaxation to provide evidence for a molecular contact surface between ice and AFP III at moderate freezing temperatures of −35°C. This phenomenon is potentially unique for AFPs because ubiquitin shows no such cross relaxation or cross saturation with ice. On the other hand, we detected liquid hydration water and strong water-AFP III and water-ubiquitin cross peaks in frozen solution using relaxation filtered 2 H and HETCOR spectra with additional 1 H-1 H mixing. These results are consistent with the idea that ubiquitin is surrounded by a hydration shell, which separates it from the bulk ice. For AFP III, the water cross peaks indicate that only a portion of its hydration shell (i.e., at the ice-binding surface) is in contact with the ice lattice. The rest of AFP III's hydration shell behaves similarly to the hydration shell of non-ice-interacting proteins such as ubiquitin and does not freeze together with the bulk water.hydration shell | type III AFP | ice binding | water solute interaction A ntifreeze proteins (AFPs) can be found in a variety of coldadapted organisms including bacteria, plants, insects, and fish (1). AFPs lower the freezing point of a given solution below its melting point, a process called thermal hysteresis. This thermal hysteresis is thought to result from the strong binding of the AFP to the surface of ice crystals making their further growth energetically unfavorable (2, 3). One of the most intensely studied classes of AFPs is the type III AFPs from the ocean pound. In particular the HPLC-12 isoform (in the following just termed AFP III), was studied with X-ray crystallography (4), NMR (5, 6), mutagenesis (4,7,8), and molecular dynamics (9). These studies showed that the ice-binding site of AFP III is a flat surface formed by predominantly nonpolar residues (5, 8), which is remarkable because the solvent accessible surface of other soluble proteins like ubiquitin are predominantly polar. There have been limited opportunities to study the water molecules at the ice-binding surface of AFPs, although recent computational and sub-angstrom X-ray structures indicate ordered water molecules at the ice-binding surface (9, 10).Ice, with its very slow 1 H spin-lattice relaxation rate (R 1 ) and very fast spin-spin relaxation rate (R 2 ), is relatively difficult to study with NMR. Measurements of relaxation rates of ice showed that R 2 is on the order of 10 6 s −1 and R 1 on the order of 10at a temperatures of about 260 K...

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mentioning

confidence: 64%

Protein–ice interaction of an antifreeze protein observed with solid-state NMR

Siemer

Huang

McDermott

2010

Proc. Natl. Acad. Sci. U.S.A.

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“…Solid state NMR spectroscopy under magic angle spinning is a method of choice for the characterization of large protein systems [114,115]. For the application to chaperone-substrate systems, ssNMR may be a useful method, when the complexes are long-lived.…”

Section: Solid State Nmr Spectroscopymentioning

confidence: 99%

Chaperones and chaperone–substrate complexes: Dynamic playgrounds for NMR spectroscopists

Burmann¹,

Hiller²

2015

Progress in Nuclear Magnetic Resonance Spectroscopy

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“…The tumbling correlation time and thus the molecular weight of the protein is of no importance, as internal dynamics, which are responsible for relaxation, are independent of the size of the molecule. This allows to achieve narrow lines for large proteins in the crystalline state (Tian et al 2009) as well as in tumbling impaired solutions (Mainz et al 2009). Using highly deuterated proteins, we obtain line widths for 1 H and 15 N which are on the order of 20 and 10 Hz, respectively (Chevelkov et al 2006).…”

Section: Introductionmentioning

confidence: 99%

Narrow carbonyl resonances in proton-diluted proteins facilitate NMR assignments in the solid-state

2010

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HNCO/HNCACO type correlation experiments are an alternative for assignment of backbone resonances in extensively deuterated proteins in the solid-state, given the fact that line widths on the order of 14-17 Hz are achieved in the carbonyl dimension without the need of high power decoupling. The achieved resolution demonstrates that MAS solid-state NMR on extensively deuterated proteins is able to compete with solution-state NMR spectroscopy if proteins are investigated with correlation times s c that exceed 25 ns.

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Large Protein Complexes with Extreme Rotational Correlation Times Investigated in Solution by Magic-Angle-Spinning NMR Spectroscopy

Cited by 89 publications

References 21 publications

Protein–ice interaction of an antifreeze protein observed with solid-state NMR

Protein–ice interaction of an antifreeze protein observed with solid-state NMR

Chaperones and chaperone–substrate complexes: Dynamic playgrounds for NMR spectroscopists

Narrow carbonyl resonances in proton-diluted proteins facilitate NMR assignments in the solid-state

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