Diffusion ordered NMR spectroscopy (DOSY) can be used to determine the apparent molecular weight of proteins (Mprotein) by measuring self-diffusion coefficient (Dt-protein). However, Dt-protein is also affected by the property of buffer solution, making the measurements complicated. In this work, 3-(trimethylsilyl) propane sodium sulfonate (DSS) was used as the internal reference molecule and the diffusion coefficient ratio between the target protein and DSS (Dr) was used to characterize the protein in solution. This method reduced the effects of buffer system on Mprotein determination, yielding more accurate Mprotein measurement. The Dr values of reference proteins with different molecular weight were determined. A correlation curve between Dr and Mprotein was established (lgMprotein=-2.6488 lgDr-0.7863, R2=0.997). Finally, Dr of the C-terminal domain of SARS-CoV main protease (Mpro-C) was determined and to yield its molecular weight. Accuracy and practicability of the fitting curve were demonstrated.
LI Hong-wei
,
YUAN Zhi-liang
,
XIA Bin
. Determination of Apparent Protein Molecular Weight in Solution by Diffusion Ordered NMR Spectroscopy[J]. Chinese Journal of Magnetic Resonance, 2018
, 35(3)
: 280
-286
.
DOI: 10.11938/cjmr20182625
[1] DOIG A J, DERREUMAUX P. Inhibition of protein aggregation and amyloid formation by small molecules[J]. Curr Opin Struct Biol, 2015, 30:50-56.
[2] YANG-HARTWICH Y, SOTERAS M G, LIN Z P, et al. p53 protein aggregation promotes platinum resistance in ovarian cancer[J]. Oncogene, 2015, 34(27):3605-3616.
[3] CHEN Y H, ZHANG Z T, BAI J, et al. Inhibition mechanisms of protein disulfide isomerase on a-synuclein Fibril aggregation[J].Chinese J Magn Reson, 2017, 34(2):131-136. 陈艳华, 张则婷, 白佳, 等. PDI抑制α-synuclein纤维化聚集作用机制研究[J]. 波谱学杂志, 2017, 34(2):131-136.
[4] JOHNSON JR C S. Diffusion ordered nuclear magnetic resonance spectroscopy:principles and applications[J]. Prog NMR Spect, 1999, 34:203-256.
[5] KE E F, CHEN Z, YE J L, et al. 2D diffusion-ordered nuclear magnetic resonance spectroscopy for structure analysis of mixtures[J]. Journal of Xiamen University-Natural Science, 1999, 38(3):366-370. 柯恩烽, 陈忠, 叶剑良, 等. 混合物溶液结构的二维扩散排序核磁共振谱[J]. 厦门大学学报(自然科学版), 1999, 38(3):366-370.
[6] DEHNER A, KESSLER H. Diffusion NMR spectroscopy:folding and aggregation of domains in p53[J]. Chem Bio Chem, 2005, 6(9):1550-1565.
[7] MACCHIONI A, CIANCALEONI G, ZUCCACCIA C, et al. Determining accurate molecular sizes in solution through NMR diffusion spectroscopy[J]. Chem Soc Rev, 2008, 37(3):479-489.
[8] FLOQUET S, BRUN S, LEMONNIER J F, et al. Molecular weights of cyclic and hollow clusters measured by DOSY NMR spectroscopy[J]. J Am Chem Soc, 2009, 131(47):17254-17259.
[9] MALLOL R, RODRÍGUEZ M A, HERAS M, et al. Particle size measurement of lipoprotein fractions using diffusion-ordered NMR spectroscopy[J]. Anal Bioanal Chem., 2012, 402(7):2407-2415.
[10] ZHANG F, YU X Y, CHEN Z, et al. Characterizations of some N-substituted-salicylhydrazide in mixtures by NMR diffusion ordered spectroscopy[J]. Chinese J Struct Chem, 2003, 22(3):287-292.
[11] MANSFIELD S L, JAYAWICKRAMA D A, TIMMONS J S, et al. Measurement of peptide aggregation with pulsed-field gradient nuclear magnetic resonance spectroscopy[J]. Biochim Biophys Acta, 1998, 1382(2):257-265.
[12] YAO S, HOWLETT G J, NORDON R S. Peptide self-association in aqueous trifluoroethanol monitored by pulsed field gradient NMR diffusion measurements[J]. J Biomol NMR, 2000, 16(2):109-119.
[13] CHEN C G, ZHANG M M, LIU Y P. Study on the Interaction between β-Cyclodextrin and naproxen by two-dimensional diffusion ordered NMR Spectroscopy[J]. J Instrum Anal, 2011, 30(9):959-963. 陈昌国, 张明明, 刘渝萍. 二维扩散排序核磁共振谱法研究β-环糊精/萘普生的相互作用[J]. 分析测试学报, 2011, 30(9):959-963.
[14] PRICE W S, NARA M, ARATA Y. A pulsed field gradient NMR study of the aggregation and hydration of parvalbumin[J]. Biophys Chem, 1997, 65(2, 3):179-187.
[15] LIN M, LARIVE C K. Detection of insulin aggregates with pulsed-field gradient nuclear magnetic resonance spectroscopy[J]. Anal Biochem, 1995, 229(2):214-220.
[16] AUGEx S, SCHMIT P, CRUTCHFIELD C A, et al. NMR measure of translational diffusion and fractal dimension. Application to molecular mass measurement[J]. J Phys Chem B, 2009, 113(7):1914-1918.
[17] WANG L M, QIU R C, HUANG S H. Quantitative analysis of active ingredients in compound acetylsalicylic acid tablets by DOSY[J]. Chinese J Magn Reson, 2016, 33(3):415-421. 王丽敏, 仇汝臣, 黄少华. 复方乙酰水杨酸片中有效成分的DOSY技术分析[J]. 波谱学杂志, 2016, 33(3):415-421.
[18] ZHONG N, ZHANG S N, ZOU P, et al. Without its N-finger, the main protease of severe acute respiratory syndrome coronavirus can form a novel dimer through its C-terminal domain[J]. J Virol, 2008, 82(9):4227-4234.
