Dmso d6

All chemical reagents were ACS grade or higher unless otherwise indicated. All buffers were passed through Chelex-100 (Bio-Rad, Hercules, CA) to remove trace metals. The D₂O, d₆-DMSO, ¹⁵NH₄Cl, and ¹³C-labeled glucose were purchased from Cambridge Isotope Laboratories, Inc. (Andover, MA).

All chemical reagents were American Chemical Society grade or higher unless otherwise indicated. The D₂O, D₆-DMSO, ¹⁵NH₄Cl, and ¹³C-labeled glucose were purchased from Cambridge Isotope Laboratories, Inc. (Andover, MA). All buffers were passed through and/or stored with dialysis bags containing Chelex-100 (Bio-Rad, Hercules, CA) to remove trace metals. The Chelex-100 treated reagents were stored in plastic containers, transferred minimally, and contacted only plastic or quartz cuvettes that had been acid washed to minimize contaminating calcium.

Most reagents were used without further purification including HB (Sigma-Aldrich, ≥99.5%), 2,2,4-trimethylpentane (isooctane, Sigma-Aldrich, ≥99.0%), deuterium oxide (D₂O, Cambridge Isotope Laboratories, 99.9%), 1,2-dipalmitoyl-sn-glycero-3phosphocholine (DPPC, Avanti Polar Lipids Inc., >99%), d₆-dimethyl sulfoxide containing tetramethyl silane (d₆-DMSO, Cambridge Isotope Laboratories, 99.9% + 0.05% TMS), activated charcoal (Sigma-Aldrich, 8−20 mesh), methanol (Omnisolve, 99.9%), 3-(trimethylsilyl)propane-1-sulfonic acid (DSS, Wilmad,), hexane (Fisher Scientific, 99.9%), and isopropanol (EMD, 99.8%). Bis(2-ethylhexyl)sulfosuccinate sodium salt (AOT, Aldrich, 99.8%) was purified using activated charcoal and methanol to remove acidic impurities as described previously.^{42 (link)} All pH measurements were conducted using a Thermo Orion 2 Star pH meter with a VWR semimicro pH probe. When conducting the NMR experiments, deuterium oxide was used in the presence of aqueous solutions and the pH was adjusted to consider the presence of deuterium (pD = 0.4 + pH).^{42 (link)} The pD is customarily referred to as pH and therefore we refer to pD as pH.^{42 (link)}

All 2D-NMR data were acquired at 27 °C on a Bruker Avance DRX 500 MHz spectrometer equipped with a QCI CryoProbe and a B-ACS 60-slot autosampler. For the 2D-HSQC experiments, two constructs were used: (i) selectively [¹³C-methyl]-methionine labeled wild-type KSHV Pr; and (ii) uniform ¹⁵N/¹H and selectively [¹³C-methyl]-isoleucine labeled KSHV Pr Δ196. Nominal protein and fragment concentrations were 20 – 25 µM and 490 – 500 µM, respectively. All other protein sample conditions, data acquisition parameters and analysis were as previously described.^{[21 (link)–23 (link)]}1D-STD NMR experiments were acquired at 27 °C on a Bruker Avance AV 800 MHz Spectrometer equipped with a TXI CryoProbe. Unlabeled wild-type HCMV Pr was diluted to 10 µ;M in deuterated buffer containing 25 mM potassium phosphate pH 8.0, 150 mM KCl, 0.1 mM EDTA, and 1 mM TCEP. Fragments were added to a final concentration of 500 µ;M from 50 mM stocks diluted in d₆-DMSO (Cambridge Isotope Laboratories, Inc.). Final sample volumes were 0.45 mL. Data were acquired using a saturation-transfer difference program equipped with a Watergate solvent suppression pulse sequence.^{[33 (link)]} On-resonance experiments were set to 0.9 ppm, while off-resonance experiments were set to 30 kHz from the spectral offset. 1D-STD NMR data was analyzed using Mnova NMR (Mestrelab Research) or ACD NMR Processor (ACD/Labs).