Sequencing Methodology

In order to overcome some solubility problems, Garrick and Sloan have suggested that small peptides may be sequenced in the spinning cup type of automatic sequencer after covalent attachment to cytochrome c. The acetylated A terminus of the cytochrome minimizes the risk of confusion due to its own 

A variation on the use of aminopeptidases to obtain information about A -terminal sequences has been reported in connection with work on spinach plastocyanin. Dipeptidyl aminopeptidases were used to cleave the A-terminal portion giving rise to a number of dipeptides which were analysed, allowing the sequence to be established.  

Methods for sequence determination by sequential d radation from the C-terminal end have in neral not been well developed. In an attempt to improve this situation. Louden and co-workers have investigated various chemical methods for determining C-terminal residues and degrading peptides from the C-terminal end. -  

Identification by gas-liquid chromatography of the thiohydantoins which result from ammonium thiocyanate degradation has been discussed.  

Mass spectrometry (electron impact, field desorption, and chemical ionization ) in principal provides either a completely different, alternative method for peptide sequencing or a valuable supplement to the standard method. Applications have, however, been relatively few so far. Further studies on the mass spectral patterns obtained from model compounds have been reported. These should allow inter alia distinctions to be made between a- and y-glutamyl peptides and between leucyl and isoleucyl residues. Sequence studies on myoglobin from the bottle-nosed dolphin were facilitated by a computer comparison method. In this method, a peptide was either deuteroacetylated and deuteropermethylated or subjected to one step of the Edman degradation, followed by deuteroacetylation and deuteropermethylation. Comparison of the mass spectra of these derivatives aided the analysis.  

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Most systems use a time-sequence to control the cleaning frequency. If the particulate load entering the bag-house is constant, this approach would be valid. However, the incoming load generally changes constantly. As a result, the straight time-sequence methodology does not provide the most efficient mode of operation. 

Premaratne, S., Mandel, M. Mower, H.F. (1995) Detection of mutagen specific adduct formation in DNA using sequencing methodology. Int. J. Biochem. Cell Biol., 27, 789-794 Quintero-Ruiz, A., Paz-Neri, L.L. Villa-Trevino, S. (1981) Indirect alkylation of CBA mouse... 

The most important recent advance of the solution NMR pulse sequence methodology is transverse relaxation optimized spectroscopy (TROSY) due to its significant sensitivity enhancement for large proteins by increasing the lifetimes of NMR signals.17 120 124-126... 

Furocoumarin photoaddition takes place predominantly with thymine. The sequence specificity has been investigated using DNA sequencing methodology. The results revealed that thymines in a GC environment have weak reactivity, while adjacent thymines are better targets. In addition, there is a strong preference for 5 -TpA sites compared with 5 -ApT, and (AT)n sequences are the most reactive towards photoaddition... 

The crustaceans, a very diverse and ancient group of arthropods, have been largely studied in order to understand animal evolution and physiology, and for use as models in environmental studies. Advances in sequencing methodologies have resulted in a significant volume of information related to genomics. For an in-depth and comprehensive review see Stillman et al. (2008). 

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