Peptide mass fingerprinting is one of the most used method for identification of protein composition and structure.
Mass spectrometry is used to precisely calculate the absolute masses of proteins which are initially cut to obtain smaller peptides. Spectrometry techniques used are MALDI-TOF (Matrix Assisted Laser Desorption/Ionization -Time of Flight) or ESI-TOF (Electrospray Ionization Time of Flight).
The peptide masses are then connected either to a database comprising known protein sequences or even the genome. This is accomplished in following steps:
- Using computer program that can translate the known genome of the organism into proteins,
- Slicing of proteins into smaller peptides of to make the process easy
- Peptidesobtained from each protein are assayed and their absolute mass is calculated
- Absolute mass obtained in the previous step is compared with the theoretical masses of peptides which are coded by C.
- The outcomes are statistically examined to get the closest match amid the two.
The advantages of this technique are:
- Genome transformed peptide mass databases can be taken as reference for mass determination.
- Inefficient de novo peptide sequencing is then needless.
The disadvantages of this technique are:
- The protein sequence has to be present in the database.
- Furthermore, most peptide mass fingerprinting algorithms undertake that the peptides come from a single protein which can complicate the result to a great extent if a mixture is present.
- Mixtures beyond a number of two to three proteins mainly need the supplementary use to accomplish appropriate specificity of identification.
The development of MALDI-MS and ESI-MS and the swift growth of databases have transformed the identification of proteins. Peptide mass fingerprinting by MALDI-MS is the vital method for high throughput identification of well-separated proteins. Mass accuracy, automatic measurement and sensitivity have been improved prominently for MALDI-TOF-MS, smoothing protein identification by Peptide mass fingerprinting with high confidence