What is the proteome?
The proteome is the mechanistic expression of gene expression, representing all proteins present in a cell or system at a given time and condition.
Why is the proteome considered dynamic?
Because protein expression varies between cells, over time, and in response to environmental or biological conditions.
How many primary amino acids are used in protein synthesis?
20 primary amino acids are used for protein synthesis in all life forms.
What are isobaric amino acids?
Amino acids with the same molecular formula and mass but different structures, e.g. leucine and isoleucine.
Why is exact mass used in proteomics instead of average molecular weight?
Exact mass allows accurate identification of peptides using high-resolution mass spectrometry.
What is mass resolution in mass spectrometry?
The ability to distinguish between closely related mass peaks; higher resolution means better mass accuracy.
What does scan speed refer to in mass spectrometry?
How fast the mass analyzer scans the mass range; higher scan speed allows faster data acquisition.
What is shotgun (bottom-up) proteomics?
A proteomics approach where proteins are digested into peptides before LC–MS/MS analysis.
What is the first step in bottom-up proteomics sample preparation?
Protein extraction from biological samples using lysis buffers.
Why is protein quantification important before digestion?
To ensure optimal trypsin-to-protein ratio and efficient digestion.
What is the role of SDS-PAGE in proteomics?
To fractionate proteins by molecular weight and reduce sample complexity.
What happens during protein digestion in bottom-up proteomics?
Proteins are enzymatically cleaved into peptides, usually using trypsin.
Why is sample clean-up required before LC–MS analysis?
To remove salts and contaminants and improve MS sensitivity.
What is the Bradford assay used for?
Estimating protein concentration using a dye-binding colorimetric method.
Why are calibration curves used in protein estimation?
To calculate unknown protein concentrations from absorbance values.
What is MS1 in mass spectrometry?
The stage where intact peptide masses (m/z) are measured.
What is MS2 used for?
Fragmenting selected ions to obtain structural information for peptide identification.
What are b-ions and y-ions?
Fragment ions generated during peptide fragmentation; b-ions from N-terminus and y-ions from C-terminus.
What is MALDI?
Matrix-Assisted Laser Desorption/Ionization, a technique for analyzing intact proteins without chromatography.
How does MALDI differ from bottom-up proteomics?
MALDI analyzes intact proteins directly, whereas bottom-up proteomics analyzes digested peptides.
What is top-down proteomics?
A proteomics approach that studies intact proteins rather than peptide fragments.
What are post-translational modifications (PTMs)?
Chemical modifications to proteins after synthesis that regulate function, activity, and localization.
Name common types of PTMs
Phosphorylation, glycosylation, ubiquitination, acetylation, methylation, lipidation, proteolysis.
Why are PTMs biologically important?
They regulate protein activity, stability, interactions, and cellular localization.
