Example ZNova Parameter Files
In this section we will provide you with a few example ZNova parameter files that should help you to get going with your applications. If you download any of the parameter files make sure to customize them for your own applications by selecting the appropriate input and output mass ranges, results directory, and logo file.
Tryptic Peptide Analysis:
Click here to download peptide.params.
This parameter file is useful for processing tryptic peptide LC/MS data. Note how we have set the Normalize Scores parameter to 3 in this case. We do this as we expect most of our peptides will contain 1-3 charges. We have also set the maximum deconvoluted mass to 5000 Da, as we do not expect any of our tryptic peptides to be larger than 5 kDa. We've changed the Minimum Score to 0.5 from the default value of 2. Peptides generally produce lower scores than proteins, since there are fewer peaks contributing to the charge state series.
Oligonucleotide Analysis:
Click here to download oligo.params
We have used this parameter file to process oligonucleotide LC/MS data (up to 50 bp in length). Note we have set the Adduct Ion to -1.0079 since we acquired our data in negative ion mode. We also chose Comprehensive deconvolution mode, as sometimes the mixtures created by failure sequences can lead to fairly complex ESI spectra.
Large Protein with High Baseline Noise:
Click here to download bsa.params
Below is an ESI spectrum of bovine serum albumin (BSA). Note the fairly high baseline (hump) under the spectrum. In the parameter file above we have set the Baseline Factor to 0.7. We have also chosen Comprehensive mode as there appears to be significant micro-heterogeneity in the sample. The mass range of the deconvolution was set to 20000 - 80000 Da, as we expect the major component to appear at 66000 Da. We also adjusted the First M/z and Last M/z to 1100 - 2000, since there are really no significant peaks below 1100 u.
You can review the results of using the parameter file above on this BSA spectrum here.
Protein with Uneven or Sloping Baseline Noise:
Click here to download 30kd.params.
Below is an example of an ESI spectrum of a 30 kD protein with a sloping baseline and very broad peaks.
We used a ZNova baseline removal factor of 0.7 to reduce the baseline as shown below.
Despite the success of baseline removal, this spectrum can still pose a problem due to the noisy part of the spectrum below m/z 1200. When ZNova determines the noise level of a spectrum, it computes the noise based on the average intensity of the input data. In most cases this approach works well if the baseline has been effectively removed. In this case, however, the noise at the left hand portion of the spectrum would adversely affect this noise calculation by over-estimating the noise level. As a result, we would most likely not observe some of the minor peaks in the deconvolution results that may be of interest. This situation can be remedied by (1) setting the input First m/z and Last m/z to the range where data are likely useful (e.g., m/z 1200 - 1800 in this case) and (2) using the Intensity Threshold parameter. The Intensity Threshold allows you to over-ride the automatic calculation of noise and enter a noise level that is based on the % relative intensity of the base peak in the spectrum. In this case we chose a value of 1 (for 1% relative intensity). We also set the peak width and merge width to 10 and 1, respectively, to deal with the very broad peaks in the input spectrum. You can view the results of this deconvolution by clicking here. The parameter file that we used may be download here.
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