MALDI data interpretation

The presence of three polypeptides in Table 5.8 tliat were not predicted from the relationship between the amino acid sequence and the enzyme used for digestion is worthy of note when interpretation of data of this sort is undertaken. The MALDI data showed six further unexpected polypeptides, none of which were detected in the LC-MS data ... 

This chapter describes and discusses the mostly used IMS technology—MALDI-TOF-IMS, including sample preparation, data interpretation/handling, and its applications. 

MALDI is widely used for glycan profiling due to the ease of sample preparation, potential for automation, speed of data acquisition, and the simplicity of data interpretation (i.e., only singly charged ions observed)... 

In order to interpret the resulting data, a model is developed for copolymers obtained by SEC fractionation, taking into account the fractionation conditions and specifically the number of fractions. The model predicts the composition and the ratio D(x) of the SEC fraction. D(x) is the ratio between the number-average and the weight-average molar mass, and x is the fraction number. The predictions of the model are compared with SEC-NMR and SEC-MALDI data for the random copolymer of styrene and MMA reacted at high conversion. 

Here, we will describe a range of applications of MALDI-MS, from the concepts of in-depth analysis of purified proteins to applications of MALDI-MS in a broader, proteomics-based research where proteins are identified, characterized, and quantified. In addihon, issues of sample preparation, protein characterization and identification strategies and bioinformatic tools for data interpretation wiU be discussed. The concepts of peptide fragmentation, sequencing and derivatization, analysis of post-translational modifications and the clinical apphcations of MALDI-MS are also briefly outlined. 

Spinali S, van Belkum A, Goering RV, Girard V, Welker M, Van Nuenen M, Pincus DH, Arsac M, Durand G. Microbial typing by MALDI-TOF MS Do we need guidance for data interpretation J Clin Microbiol. 2014. doi 10.1128/jcm.01635-14. 

Comparing the mass spectra of the interaction experiments with those acquired from control experiments reveals differences that can be related to the structure of the interaction complex. Depending on the reaction, intact proteins, proteolytic peptides or both are simultaneously analyzed by MS and, optionally, MS/MS. If the sample is too complex for direct analysis, a broad range of additional means of separation are available, e.g., electrophoresis, LC, or affinity capture, which can all be efficiently combined with ESI or MALDI MS (Sects. 4.7 and 4.8). A major challenge for all three techniques described below is the data interpretation. This concerns less the identity of the resulting products than the molecular puzzle they create. Relating the observed differences between sample and control experiments to the structure of the interaction complex can be difficult, and great care is recommended before the data in hand are considered evidence for the existence of a certain structural element. 

The nature of the termination reaction in MMA polymerization has been investigated by a number of groups using a wide range of techniques (Tabic 5.5), There is general agreement that there is substantial disproportionation. However, there is considerable discrepancy in the precise values of k tk. In some cases the difference has been attributed to variations in the way molecular weight data are interpreted or to the failure to allow for other modes of termination under the polymerization conditions (chain transfer, primary radical termination).154 In other eases the reasons for the discrepancies are less clear. MALDI-TOF mass... 

Experimentation showed that the protein was not glycosylated and that the sequence at the iV-amino acid terminus corresponded to that expected. The C-terminus sequence, however, did not correspond to that predicted and these data were interpreted in terms of the presence of a heterogeneous, truncated, protein. A study of the tryptic digest fragments from this protein with matrix-assisted laser desorption ionization (MALDI) with post-source decay enabled the authors to suggest the positions at which the parent protein had been truncated. 

The final step in any proteomic study is the identification of proteins using the MS data. This is highly dependent on large databases of protein sequences and computer algorithms that can compare and interpret the MS spectra using these databases. Since the output of MALDI-TOF and ESTMS-MS analyses are essentially different, they employ different types of computer software for the interpretation of MS spectra. 

Peptide fragments from PSD and CAD processes are usually formed via peptide bond cleavages (i.e., between the backbone NH of one residue and the carbonyl of the adjacent residue). Many of the same principles apply for interpretation of ESI-MS/MS data as for MALDI-PSD. When the charge... 

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