A Solid-State NMR Study of Protein Hydration and Stability

Pharmaceutical Research - Tập 15 Số 12 - Trang 1816-1821 - 1998
Separovic, F.1, Lam, Y. H.1, Ke, X.2, Chan, H.-K.2
1School of Chemistry, University of Melbourne, Parkville, Australia
2Department of Pharmacy, University of Sydney, Sydney, Australia

Tóm tắt

Purpose. The mobility of protein in powders at different hydration levels was studied in relation to aggregation and activity. Methods. Magic angle spinning 13C, 15N, 1H, 2H, and 17O NMR techniques were used to determine changes in the mobility of surface residues in proteins as a function of hydration and related to changes in activity. NMR relaxation measurements of high frequency (ω0, T1) and low frequency (ω1, T1p) motions have been carried out on lyophilized DNase, insulin and lysozyme stored at different relative humidities. Moisture-induced aggregation and enzymatic activity of the lyophilized proteins was determined by high performance size exclusion chromatography and bioassays. Results. There was little change in T1p observed with increasing humidity. The results show, however, that there is a decrease in T1, for DNase, insulin and lysozyme at relative humidities ranging from 0−98%, and we propose that the reduction in T1, is related to the aggregation susceptibility of proteins during storage at different humidities. The water mobility was determined directly using 17O NMR experiments. We found that as the amount of weakly-bound water increases, the protein surface mobility decreases and is coupled with increased aggregation. Aggregation measurements at different humidities were correlated with bioassays for lysozyme and found to be consistent with the hydration data. Conclusions. Mobility of protein molecules was determined by solid-state NMR over a wide range of % RH and it was found that water content leads to a change in mobility of protein molecules. The aggregation and activity of proteins were strongly correlated to change in molecular mobility.

Từ khóa


Tài liệu tham khảo

citation_journal_title=Science; citation_title=Drug delivery takes a deep breath; citation_author=R. F. Service; citation_volume=277; citation_publication_date=1997; citation_pages=1199-1200; citation_id=CR1

citation_title=Stability of Protein Pharmaceuticals Part A Chemical and Physical Pathways of Protein Degradation; citation_publication_date=1992; citation_id=CR2; citation_author=T. J. Ahern; citation_author=M. C. Manning; citation_publisher=Plenum Press

citation_journal_title=Biopolymers; citation_title=The influence of hydration on the conformation of lysozyme studied by solid state 13C-NMR spectroscopy; citation_author=R. B. Gregory, M. Gangoda, R. K. Gilpin, W. Su; citation_volume=33; citation_publication_date=1993; citation_pages=513-519; citation_id=CR3

citation_journal_title=Pharm. Res.; citation_title=Moisture-induced aggregation of lyophilized insulin; citation_author=H. R. Costantino, R. Langer, A. M. Klibanov; citation_volume=11; citation_publication_date=1994; citation_pages=21-29; citation_id=CR4

citation_journal_title=Pharm. Res.; citation_title=Determination of molecular mobility of lyophilized Bovine Serum Albumin and γ-Globulin by solid-state H NMR and relation to aggregation-susceptibility; citation_author=S. Yoshioka, Y. Aso, S. Kojima; citation_volume=13; citation_publication_date=1996; citation_pages=926-930; citation_id=CR5

R. K. Harris. State of the art for solids. Chemistry in Britain, pp. 601–604 (1993).

C. R. Kahn and L. C. Harrison, Receptor Biochemistry and Methodology, volume 12 A. Methods for study of structure and functions, Liss, New York, 1988.

citation_title=Lysozymes: Model Enzymes in Biochemistry and Biology; citation_publication_date=1996; citation_id=CR8; citation_author=P. Jolles; citation_publisher=Birkhauser

citation_journal_title=Pharm. Res.; citation_title=Spray dried powders and powder blends of recombinant human deoxyribonuclease (rhDNase) for aerosol delivery; citation_author=H.-K. Chan, A. Clark, I. Gonda, M. Mumenthaler, C. Hsu; citation_volume=14; citation_publication_date=1997; citation_pages=431-437; citation_id=CR9

citation_title=Handbook of Chemistry and Physics; citation_publication_date=1996; citation_id=CR10; citation_publisher=CRC Press

citation_journal_title=Drug Dev. Ind. Pharm.; citation_title=A standard method for determination of lysozyme activity via controlling the kinetics of substrate-enzyme interactions; citation_author=A. E. El-Nimr; citation_volume=11; citation_publication_date=1985; citation_pages=653-662; citation_id=CR11

citation_title=Analytical NMR; citation_publication_date=1989; citation_id=CR12; citation_author=L. D. Field; citation_author=S. Sternhell; citation_publisher=Chichester Press

citation_title=High Resolution NMR, Theory and Chemical Application; citation_publication_date=1980; citation_id=CR13; citation_author=E. D. Becker; citation_publisher=Academic Press

citation_journal_title=J. Chemical Education; citation_title=Spin-lattice relaxation times in 1H NMR spectroscopy; citation_author=D.J. Wink; citation_volume=66; citation_publication_date=1989; citation_pages=810-813; citation_id=CR14

The Merck Index, 10th edition, An Encyclopedia of Chemicals, Drugs and Biologicals, Merck & Co. Inc., Rahway NJ, USA (1983).

citation_journal_title=Chem. Phys. Lett.; citation_title=Cross relaxation effects on the longitudinal relaxation rate of water in protein solutions; citation_author=G. Valensin, N. Niccolai; citation_volume=79; citation_publication_date=1981; citation_pages=47-50; citation_id=CR16

citation_title=Fourier Transform NMR Spectroscopy; citation_publication_date=1976; citation_id=CR17; citation_author=D. Shaw; citation_publisher=Elsevier Scientific Pub. Co.

citation_title=High Resolution Nuclear Magnetic Resonance Spectroscopy; citation_publication_date=1965; citation_id=CR18; citation_author=J. W. Emsley; citation_author=J. Feeney; citation_author=L. H. Sutcliffe; citation_publisher=Pergamon Press

citation_journal_title=J. Phys. Chem.; citation_title=The spin-lattice relaxation of oxygen-17 in water; citation_author=J. C. Hindman, A. Svirmickas, M. Wood; citation_volume=74; citation_publication_date=1970; citation_pages=12; citation_id=CR19

citation_journal_title=Biophys. J.; citation_title=Nuclear magnetic resonance evidence using D2O for structured water in muscle and brain; citation_author=F. W. Cope; citation_volume=9; citation_publication_date=1969; citation_pages=303-319; citation_id=CR20

citation_journal_title=Methods in Enzymology; citation_title=Dynamic properties of water at phosphatidylcholine lipid-bilayer surfaces as seen by deuterium nuclear magnetic resonances approach; citation_author=F. Volke, S. Eisenblaetter, J. Galle, G. Klose; citation_volume=220; citation_publication_date=1993; citation_pages=143-157; citation_id=CR21

citation_journal_title=Biotech. and Bioengineer.; citation_title=Moisture-induced aggregation of lyophilized proteins in the solid state; citation_author=W. R. Liu, R. Langer, A. M. Klibanov; citation_volume=37; citation_publication_date=1991; citation_pages=177-184; citation_id=CR22

citation_journal_title=PDA J. Pharm. Sci. and Tech.; citation_title=Aggregation of insulin and its prevention by carbohydrate excipients; citation_author=M. Katakam, A. K. Banga; citation_volume=49; citation_publication_date=1995; citation_pages=160-165; citation_id=CR23

citation_journal_title=J. Pharm. Pharmacol.; citation_title=The aggregation of bovine serum albumin in solution and in solid state; citation_author=G. M. Jordan, S. Yoshioka, T. Terao; citation_volume=46; citation_publication_date=1994; citation_pages=182-185; citation_id=CR24

citation_journal_title=Biophys. J.; citation_title=Dehydration-induced comformational transitions in proteins and their inhibition by stabilizers; citation_author=S. J. Prestrelski, N. Tedeschi, T. Arakawa, J. F. Carpenter; citation_volume=65; citation_issue=2; citation_publication_date=1993; citation_pages=661-671; citation_id=CR25

citation_journal_title=Proc. Nat. Acad. Sci. USA; citation_title=Lyophilization-induced reversible channels in the secondary structure of proteins; citation_author=K. Griebenow, A. M. Kibanov; citation_volume=92; citation_publication_date=1995; citation_pages=10969-10976; citation_id=CR26

Thông tin
Thông tin xuất bản

Pharmaceutical Research

Tập 15 Số 12

1816-1821

Thông tin tác giả