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Proteins with no missing values in three replicates were accepted as identified proteins for the protein quantification analysis.
This included for example the sequence coverage, score, rank and molecular weight of identified proteins for each spot on the gel in question (Figure 3C).
To view the bait-enriched protein complexes/networks, the identified proteins for each bait protein were organized by their major function.
We applied yeast two-hybrid and in vitro binding assays as a proof-of-concept trial for analysis of the newly identified proteins for binding to Hsp90.
Therefore we investigated those identified proteins for which the most N-terminal (tryptic/semi-tryptic and/or modified) peptide matched the predicted lTP cleavage site or was within 100 residues downstream of the predicted lTP, without any additional peptides matching upstream of the predicted lTP site.
First, we used data from all 34 identified proteins for model development.
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Different spots yielded the same identified protein for 10 proteins, with a maximum of six spots for AAEL005798 (V-ATP synthase subunit beta).
These trees were used to identify proteins for which the nearest relative was not from the Geobacteraceae.
Unlike 2D PAGE and HPLC-MS/MS, large-scale western blotting only identifies proteins for which antibodies are already available.
Literature knowledge and MLDM layers that are more easily measured than proteins, such as mRNA or genomic information, could help to identify proteins for such targeted analyses [ 62].
The same method was used to identify proteins for which the 1-year to baseline change in concentration differed between E+P and E-alone.
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