Subject(s)
Biological Therapy , Biotechnology/history , Chemistry, Pharmaceutical/history , Phytotherapy , Polyketides/therapeutic use , Propiolactone/therapeutic use , Bacteria , Fungi , History, 20th Century , History, 21st Century , Humans , Italy , Male , Plants , Polyketides/chemistry , Propiolactone/analogs & derivatives , Propiolactone/chemistryABSTRACT
Inactivation of intact influenza viruses using formaldehyde or ß-propiolactone (BPL) is essential for vaccine production and safety. The extent of chemical modifications of such reagents on viral proteins needs to be extensively investigated to better control the reactions and quality of vaccines. We have evaluated the effect of BPL inactivation on two candidate re-assortant vaccines (NIBRG-121xp and NYMC-X181A) derived from A/California/07/2009 pandemic influenza viruses using high-resolution FT-ICR MS-based proteomic approaches. We report here an ultra performance LC MS/MS method for determining full-length protein sequences of hemagglutinin and neuraminidase through protein delipidation, various enzymatic digestions, and subsequent mass spectrometric analyses of the proteolytic peptides. We also demonstrate the ability to reliably identify hundreds of unique sites modified by propiolactone on the surface of glycoprotein antigens. The location of these modifications correlated with changes to protein folding, conformation, and stability, but demonstrated no effect on protein disulfide linkages. In some cases, these modifications resulted in suppression of protein function, an effect that correlated with the degree of change of the modified amino acids' side chain length and polarity.
Subject(s)
Influenza Vaccines/chemistry , Neuraminidase/chemistry , Propiolactone/chemistry , RNA-Binding Proteins/chemistry , Viral Core Proteins/chemistry , Viral Proteins/chemistry , Virus Inactivation , Amino Acid Sequence , Antigens, Viral/chemistry , Cysteine/chemistry , Hemagglutinins/chemistry , Nucleocapsid Proteins , Polysaccharides/chemistry , Tandem Mass SpectrometryABSTRACT
In order to establish the theory and method for the identification of Ophiogon japonicus (Thunb.)Ker-Gawl. of traditional Chinese herbal medicines and its confusable varieties, second derivative FTIR spectroscopy was used combined with statistics. Samples were collected directly by Fourier Transform Infrared (FTIR) spectra with OMNI-Sampler. Then through converting FTIR spectra of the samples into second derivative spectra by derivative spectra software, Ophiogon japonicus (Thunb.)Ker-Gawl. could be identified from the confusable varieties with statistics. The result shows that the second derivative FTIR of Ophiogon japonicus (Thunb.)Ker-Gawl. and its confusable varieties are different, which differ greatly in 2 000-650 cm(-1) range in second derivative FTIR spectroscopy. The Ophiogon japonicus (Thunb.)Ker-Gawl. and its confusable varieties can be identified by identifying the inner layer parts of the cuticles of samples by second derivative FTIR spectroscopy with statistics directly, rapidly and accurately.