Proteins sequence identification

Protein sequencing Identification of the consecutive arrangement of all the amino acids that make up a protein. 

Bidargaddi, N.P. Chetty, M. Kamruzzaman, J. (2008). Hidden Markov models incorporating fuzzy measures and integrals for protein sequence identification and alignment. Genomics Proteomics Bioinformatics, Vol. 6, No. 2, pp. 98-110. 

Kinter, M. and Sherman, N.E., Protein Sequencing and Identification Using Tandem Mass Spectrometry, Wiley, Chichester, U.K., 2000. 

A potentially general method of identifying a probe is, first, to purify a protein of interest by chromatography (qv) or electrophoresis. Then a partial amino acid sequence of the protein is deterrnined chemically (see Amino acids). The amino acid sequence is used to predict likely short DNA sequences which direct the synthesis of the protein sequence. Because the genetic code uses redundant codons to direct the synthesis of some amino acids, the predicted probe is unlikely to be unique. The least redundant sequence of 25—30 nucleotides is synthesized chemically as a mixture. The mixed probe is used to screen the Hbrary and the identified clones further screened, either with another probe reverse-translated from the known amino acid sequence or by directly sequencing the clones. Whereas not all recombinant clones encode the protein of interest, reiterative screening allows identification of the correct DNA recombinant. 

Databases Comparison of DNA and protein sequences obtained from unknown gene with known sequences in databases can facilitate gene identification. 

Structural analysis of the two pectate lyases PelC and PelE (5, 6), demonstrated that these proteins fold in a large heHx of parallel P strands. A stack of asparagine residues parallel to the helix probably plays a role in the stabUity of this structure. Identification of the structurally conserved amino adds lead to a reaHgnment of the protein sequences (7). In addition to Erwinia extracellular pectate lyases, the multiple aHgnment indudes the Bacillus subtilis pectate lyase, Aspergillus tdger and E. carotovora pectin lyases and plant proteins. 

The ability to visualize spots on a 2D gel, while useful as a fingerprint, is not the same as protein identification. Protein sequencing by the Edman degradation technique is the classical means of determining... 

Ponting CP, Schultz J, Milpetz F, Bork P. SMART identification and annotation of domains from signalling and extracellular protein sequences. Nucleic Acids Res 1999 27[l] 229-232. 

Three human FFPE liver cases were analyzed and each yielded just under 20,000 distinct peptide sequence identifications and just over 3000 protein... 

In addition to MALDI-TOF and LC-MS/MS, SELDI-TOF-MS can also be used to determine expression profiling of various biological samples, such as serum or plasma for early detection of infection. Serum proteomic profiling assay, for instance, has been used to distinguish patients with acute SARS (severe acute respiratory syndrome) from patients with fever and influenza with 100% accuracy [16]. A major limitation of SELDI-TOF-MS, however, is that it cannot be used for direct amino acid sequence identification of the biomarker proteins, necessitating further sample fractionation and protein purification. 

Library A collection of antibodies, usually Fab or scFv fragments, in the range of 106 to 1010 and displayed on the surface of bacteriophage whose DNA gene contains a DNA sequence capable of expression as the antibody protein. Thus, identification of a single member of the library by selection can be used to generate multiple copies of the phage and sizeable amounts of the antibody protein. 

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