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Peptide mass fingerprint data were determined on a MALDI-TOF mass spectrometer (REFLEX IV, Bruker Daltonics, Bremen, Germany) in reflector mode.

For peptide mass fingerprint data, peak picking was conducted using GPS Explorer software version 3.6 (Applied Biosystems, Foster City, CA).

Peptide mass fingerprint data was analyzed against the conceptual translation using PAWS (Digilab, Holliston, MA) and peptide sequence tags derived with Peaks Studio.

Identification of proteins from peptide mass fingerprint data was performed using the MatrixScience 'Mascot' search engine [ 38], and was based on their 'Probability Based MOWSE Score' algorithm.

In a subsequent and very rapid experimental approach, we used mass fingerprint data to include further neuropeptide sequences from relatively conserved peptides (AKH-1 and sulfakinins) in the phylogenetic analyses.

A comparison of the peptide mass fingerprint data with the nLac indicated presence of fragments, not observed in the nLac, which were attributed to different pattern of glycosylation in the P. pastoris and which are likely to have contributed to the observed differences in some biochemical properties.

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The purified fraction was identified as a glutamate dehydrogenase (GDH) by peptide mass fingerprinting data.

The identification of enzyme according to the peptide mass fingerprinting data was performed by MASCOT search program in the SwissProt database (Barash and Mor 1973), after excluding the trypsin autolysis products from the control spectrum.

Fig. 3 Proteins up- or down-regulated twofold and significantly varying with water deficit (WD), Heat, and combined water deficit + heat (WD + H) stress compared with Control (at 5%) were excised from 2D gels, sequenced using MS MALDI-TOF, and identified by database searches using peptide mass fingerprinting data.

Fig. 4 Proteins up- or down-regulated twofold and significantly varying with water deficit (WD), Heat, and combined water deficit + heat (WD + H) stress compared with Control (at 5%) were excised from 2D gels, sequenced using MS MALDI-TOF, but not identified from the available databases using peptide mass fingerprinting data.

Peptide mass fingerprinting data (PMF) were analyzed using the MASCOT server (www.matrixscience.com).com

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