PL EN


Preferences help
enabled [disable] Abstract
Number of results
2014 | 61 | 2 | 349-357
Article title

Investigation of asparagine deamidation in a SOD1-based biosynthetic human insulin precursor by MALDI-TOF mass spectrometry

Content
Title variants
Languages of publication
EN
Abstracts
EN
A biosynthetic human insulin precursor displayed enhanced susceptibility to deamidation at one particular site. The present study was undertaken to monitor progress of precursor deamidation at successive manufacturing stages. MALDI-TOF/TOF MS in combination with controlled endoproteinase Glu-C and endoproteinase Asp-N proteolysis was used for rapid and unambiguous determination of deamidated residue within the investigated structure. Close inspection of isotopic distribution patterns of peptides resulting from enzymatic digestion enabled determination of distinct precursor forms occurring during the production process. Asn, Asp, isoAsp and succinimide derivatives of the amino acid at position 26 were unambiguously identified. These modifications are related to the leader peptide of a precursor encompassing amino acid sequence corresponding to that of superoxide dismutase [Cu-Zn] (SOD1 1, EC=1.15.1.1). Monitoring of precursor deamidation process at successive manufacturing stages revealed that the protein folding stage was sufficient for a prominent replacement of asparagine by aspartic and isoaspartic acid and the deamidated human insulin precursor constituted the main manufactured product. Conversion proceeded through a succinimide intermediate. Significant deamidation is associated with the presence of SNG motif and confirms results achieved previously on model peptides. Our findings highlight an essential role of the specific amino acid sequence on accelerated rate of protein deamidation. To our knowledge, this is the first time that such a dramatic change in the relative abundance of Asp and isoAsp resulting from protein deamidation process is reported.
Year
Volume
61
Issue
2
Pages
349-357
Physical description
Dates
published
2014
received
2014-01-23
revised
2014-05-11
accepted
2014-05-28
(unknown)
2014-06-16
References
  • Aswad DW (1995) Deamidation and isoaspartate formation in proteins. In Aswad DW, ed. CRC Press, Boca Raton.
  • Aswad DW (2000) Isoaspartate in peptides and proteins: formation, significance, and analysis. J Pharm Biomed Anal 21: 1129-1136.
  • Brange J (1992) Chemical stability of insulin. 4. Mechanisms and kinetics of chemical transformations in pharmaceutical formulation. Acta Pharm Nord 4: 209-222.
  • Carillon J, Rouanet JM, Cristol JP, Brion R (2013) Superoxide dismutase administration, a potential therapy against oxidative stress related diseases: several routes of supplementation and proposal of an original mechanism of action. Pharm Res 30: 2718-2728.
  • Catak S, Monard G, Aviyente V, Ruiz-Lopez MF (2009) Deamidation of asparagine residues: direct hydrolysis versus succinimide-mediated deamidation mechanisms. J Phys Chem A 113: 1111-1120.
  • Chelius D, Rehder DS, Bondarenko PV (2005) Identification and characterization of deamidation sites in the conserved regions of human immunoglobulin gamma antibodies. Anal Chem 77: 6004-6011.
  • Cournoyer JJ, Lin C, Bowman MJ, O'Connor PB (2007) Quantitating the relative abundance of isoaspartyl residues in deamidated proteins by electron capture dissociation. J Am Soc Mass Spectrom 18: 48-56.
  • Furukawa Y, O'Halloran TV (2006) Posttranslational modifications in Cu,Zn-superoxide dismutase and mutations associated with amyotrophic lateral sclerosis. Antioxid Redox Signal 8: 847-67.
  • Geiger T, Clarke S (1987) Deamidation, isomerization, and racemization at asparaginyl and aspartyl residues in peptides. J Biol Chem 262: 785-794.
  • Hayes CS, Setlow P (1997) Analysis of deamidation of small, acid-soluble spore proteins from Bacillus subtilis in vitro and in vivo. J Bacteriol 179: 6020-6027.
  • Kameoka D, Ueda T, Imoto TA (2003) A method for the detection of asparagine deamidation and aspartate isomerization of proteins by MALDI/TOF-mass spectrometry using endoproteinase Asp-N. J Biochem 134: 129-135.
  • Kang JH, Choi BJ, Kim SM (1997) Expression and characterization of recombinant human Cu, Zn-superoxide dismutase in Escherichia coli. J Biochem Mol Biol 30: 60-65.
  • Krokhin OV, Antonovici M, Ens W, Wilkins JA, Standing KG (2006) Deamidation of -Asn-Gly- sequences during sample preparation for proteomics: consequences for MALDI and HPLC-MALDI analysis. Anal Chem 78: 6645-6650.
  • Li B, Borchardt RT, Topp EM, VanderVelde D, Schowen RL (2003) Racemization of an asparagine residue during peptide deamidation. J Am Chem Soc 125: 11486-11487.
  • Li JR, Yu P (2007) Expression of Cu, Zn-superoxide dismutase gene from Saccharomyces cerevisiae in Pichia pastoris and its resistance to oxidative stress. Appl Biochem Biotechnol 136: 127-39.
  • O'Connor PB, Cournoyer JJ, Pitteri SJ, Chrisman PA, McLuckey SA (2006) Differentiation of aspartic and isoaspartic acids using electron transfer dissociation. J Am Soc Mass Spectrom 17: 15-19.
  • Oliyai C, Borchardt RT (1994) Chemical pathways of peptide degradation. VI. Effect of the primary sequence on the pathways of degradation of aspartyl residues in model hexapeptides. Pharm Res 11: 751-758.
  • Patel K, Borchardt RT (1990) Chemical pathways of peptide degradation. II. Kinetics of deamidation of an asparaginyl residue in a model hexapeptide. Pharm Res 7: 703-711.
  • Peters B, Trout BL (2006) Asparagine deamidation: pH-dependent mechanism from density functional theory. Biochemistry 45: 5384-5392.
  • Rivers J, McDonald L, Edwards IJ, Beynon RJ (2008) Asparagine deamidation and the role of higher order protein structure. J Proteome Res 7: 921-927.
  • Robinson NE, Robinson AB (2004) Deamidation of asparaginyl and glutaminyl residues in peptides and proteins. In Molecular Clocks. Althouse Press, Cave Junction OR.
  • Schaller G, et al. (2012) Human pharmacokinetics of intravenous recombinant human Cu/Zn superoxide dismutase. Int J Clin Pharmacol Ther 50: 413-417.
  • Shi Y, et al. (2013) Deamidation of asparagine to aspartate destabilizes Cu, Zn superoxide dismutase, accelerates fibrillization, and mirrors ALS-linked mutations. J Am Chem Soc 135: 15897-15908.
  • Sinha S, Zhang L, Duan S, Williams TD, Vlasak J, Ionescu R, Topp EM (2009) Effect of protein structure on deamidation rate in the Fc fragment of an IgG1 monoclonal antibody. Protein Sci 18: 1573-1584.
  • Wearne SJ, Creighton TE (1989) Effect of protein conformation on rate of deamidation: ribonuclease A. Proteins 5: 8-12.
  • Wilcox KC et al. (2009) Modifications of superoxide dismutase (SOD1) in human erythrocytes: a possible role in amyotrophic lateral sclerosis. J Biol Chem 284: 13940-13947.
  • Wright HT (1991) Nonenzymatic deamidation of asparaginyl and glutaminyl residues in proteins. Crit Rev Biochem Mol Biol 26: 1-52.
  • Yasui K, Baba A (2006) Therapeutic potential of superoxide dismutase (SOD) for resolution of inflammation. Inflamm Res 55: 359-63.
  • Zhang H, Joseph J, Crow J, Kalyanaraman B (2004) Mass spectral evidence for carbonate-anion radical induced posttranslational modification of tryptophan tokynurenine in human Cu,Zn superoxide dismutase. Free Radic Biol Med 37: 2018–2026
Document Type
Publication order reference
YADDA identifier
bwmeta1.element.bwnjournal-article-abpv61p349kz
Identifiers
JavaScript is turned off in your web browser. Turn it on to take full advantage of this site, then refresh the page.