By a News Reporter-Staff News Editor at Life Science Weekly -- Data detailed on Enzymes and Coenzymes have been presented. According to news reporting from Boston, Massachusetts, by NewsRx journalists, research stated, "Cystine knots or nested disulfides are structurally difficult to characterize, despite current technological advances in peptide mapping with high-resolution liquid chromatography coupled with mass spectrometry (LC-MS). In the case of recombinant human arylsulfatase A (rhASA), there is one cystine knot at the C-terminal, a pair of nested disulfides at the middle, and two out of three unpaired cysteines in the N-terminal region."
The news correspondents obtained a quote from the research from Northeastern University, "The statuses of these cysteines are critical structure attributes for rhASA function and stability that requires precise examination. We used a unique approach to determine the status and linkage of each cysteine in rhASA, which was comprised of multi-enzyme digestion strategies (from Lys-C, trypsin, Asp-N, pepsin, and PNGase F) and multi-fragmentation methods in mass spectrometry using electron transfer dissociation (ETD), collision induced dissociation (CID), and CID with MS(3) (after ETD). In addition to generating desired lengths of enzymatic peptides for effective fragmentation, the digestion pH was optimized to minimize the disulfide scrambling. The disulfide linkages, including the cystine knot and a pair of nested cysteines, unpaired cysteines, and the post-translational modification of a cysteine to formylglycine, were all determined. In the assignment, the disulfide linkages were Cys138-Cys154, Cys143-Cys150, Cys282-Cys396, Cys470-Cys482, Cys471-Cys484, and Cys475-Cys481. For the unpaired cysteines, Cys20 and Cys276 were free cysteines, and Cys51 was largely converted to formylglycine (>70%)."
According to the news reporters, the research concluded: "A successful methodology has been developed, which can be routinely used to determine these difficult-to-resolve disulfide linkages, ensuring drug function and stability."
For more information on this research see: Complete mapping of a cystine knot and nested disulfides of recombinant human arylsulfatase A by multi-enzyme digestion and LC-MS analysis using CID and ETD. Journal of the American Society for Mass Spectrometry, 2013;24(1):125-33. (Elsevier - www.elsevier.com; Journal of the American Society for Mass Spectrometry - www.elsevier.com/wps/product/cws_home/505727)
Our news journalists report that additional information may be obtained by contacting W. Ni, Barnett Institute of Chemical and Biological Analysis and Dept. of Chemistry and Chemical Biology, Northeastern University, Boston, MA 02115, United States. Additional authors for this research include M. Lin, P. Salinas, P. Savickas, S.L. Wu and B.L Karger (see also Enzymes and Coenzymes).
Keywords for this news article include: Ions, Boston, Disulfides, Hydrolases, Electrolytes, Massachusetts, United States, Arylsulfatases, Inorganic Chemicals, Enzymes and Coenzymes, North and Central America.
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