[MALD-MS in the quantitative analysis of peptides and proteins].
A modified method of isotope dilution was applied to the quantitative determination of peptides and proteins by MALDI MS at subpicomolar level. The essence of the method consists in the quantitative analysis of the enzymic hydrolysis products rather than the starting compounds. This allows the measurements to be performed at a higher resolution and makes the method independent of the molecular mass of oligopeptides and proteins examined. Fragments obtained by hydrolysis of the same oligopeptide or protein in a known concentration by the same enzyme and labeled with the stable 18O isotope are used as internal standards. The label is introduced by carrying out the hydrolysis in H(2)18O, and the oligopeptide concentration is calculated from the isotope distribution between the labeled and unlabeled hydrolysis products in the mass spectrum. This method was tested in the determination of concentrations of the angiotensinogen (1-14) fragment (oligopeptide), extracellular RNAase from Bacillus amyloliquefaciens (protein) and its protein inhibitor, barstar M. Usefulness of this method in kinetic studies was also demonstrated.
Mirgorodskaia
O A
OA
Institute of Cytology, Russian Academy of Sciences, St. Petersburg, Russia. oam@om1533.spb.edu
Koz'min
Iu P
IuP
Titov
M I
MI
Savel'eva
N V
NV
Körner
R
R
Sönksen
C
C
Miroshnikov
A I
AI
Roestorff
P
P
D019943:Amino Acid Substitution; D000808:Angiotensinogen / Q000031:analogs & derivatives* / Q000737:chemistry; D000818:Animals; D001407:Bacillus / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry; D006736:Horses; D006868:Hydrolysis; D007201:Indicator Dilution Techniques; D007700:Kinetics; D009842:Oligopeptides / Q000737:chemistry*; D010103:Oxygen Isotopes; D011506:Proteins / Q000737:chemistry*; D012260:Ribonucleases / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D001426:Bacterial Proteins; D009842:Oligopeptides; D010103:Oxygen Isotopes; D011506:Proteins; C079544:angiotensinogen (1-14); D000808:Angiotensinogen; C057438:barstar protein, Bacillus amyloliquefaciens; D012260:Ribonucleases