Protein Analysis FAQ

• • SDS-PAGE Measures Protein Molecular Weight: How Is Migration Distance Measured

  Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is a widely used technique for determining protein molecular weight. Following SDS-PAGE, the molecular weight of proteins can be estimated by measuring their migration distance within the gel. The migration distance of proteins can be determined using image analysis software (e.g., ImageJ) or a ruler by measuring the distance from the sample well to the protein band. Estimating Protein Molecular Weight Using Migration Distance: .......

• • How to Remove the SUMO Tag After Purification

  The Small Ubiquitin-like Modifier (SUMO) tag is a widely utilized fusion tag that facilitates protein solubility and stability. It can be specifically cleaved by SUMO-specific proteases, such as SENP1 or SENP2, to yield the target protein in its untagged form. The standard protocol for SUMO tag removal consists of the following steps Expression and Purification of SUMO-Tagged Protein The target protein is expressed using a SUMO-fusion expression system, commonly in Escherichia coli, and subsequently ......

• • What Is the Principle of Separating Proteins by Gel Chromatography

  Gel chromatography (also referred to as gel permeation chromatography, gel filtration chromatography, or molecular sieve chromatography) is a widely used technique for protein separation and purification. Its principle is based on differences in the molecular size and shape of proteins, which govern their interaction with the gel medium. The gel medium utilized in gel chromatography is typically composed of materials such as polyacrylamide or agarose, which form a three-dimensional network structure........

• • What Is the Essential Difference Between the Top-Down and Bottom-Up Approaches in Proteomics Analysis

  Mass spectrometry is a fundamental technology in proteomics research. In the field of protein mass spectrometry, two primary analytical approaches are employed: the Top-down and Bottom-up strategies. These approaches fundamentally differ in how they process and analyze protein samples. Top-Down Approach In the Top-down approach, intact proteins are directly introduced into the mass spectrometer and subsequently fragmented within the instrument. This means that the analysis begins with the entire protein....

• • What Is Protein Characterization

  Protein characterization encompasses the systematic analysis of the molecular weight, sequence, structure, function, interactions, and other relevant properties of protein molecules. The primary objective is to achieve a comprehensive understanding of protein properties and functions, thereby elucidating their roles in biological processes, including metabolic pathways and signal transduction. This process typically integrates a variety of experimental and computational methods to achieve a thorough .......

• • Why Does the Coloration of Thin-Layer Chromatography Appear Blurry

  Thin-layer chromatography (TLC) is a widely used technique for the separation and identification of compounds in laboratories. However, in certain cases, the visualization of spots may appear unclear. This issue can arise due to various factors, some of which are outlined below: 1. Improper Sample Application Unsteady hands or incorrect handling during sample application can result in poorly defined spots on the TLC plate. It is recommended to apply samples with precision using appropriate tools, such as...

• • Criteria for Identifying the Molecular Ion Peak in Mass Spectra

  The identification of the molecular ion peak (M⁺ peak) in mass spectra is primarily based on the following criteria: 1. Mass-to-Charge Ratio Considerations The molecular ion peak generally corresponds to the highest mass-to-charge ratio (m/z) in the spectrum, as it represents the intact molecule prior to fragmentation. 2. Nitrogen Rule According to the nitrogen rule, compounds containing an even number of nitrogen atoms tend to exhibit a molecular ion peak with an even m/z value, whereas those with an odd..

• • How to Determine if a Recombinant Protein Is a Membrane Protein

  Determining whether a recombinant protein is a membrane protein can be achieved using the following approaches: Amino Acid Sequence Analysis 1. Transmembrane Domain Prediction Bioinformatics tools such as TMHMM, Phobius, and HMMTOP can predict transmembrane domains based on the amino acid sequence of the protein. If the protein contains multiple transmembrane helices, it is likely to be a membrane protein. 2. Signal Peptide Recognition Some membrane proteins possess signal peptide sequences, which can be...

• • A Brief Description of the Advantages and Challenges of PEGylated Proteins

  PEGylation of proteins is a method that involves attaching polyethylene glycol (PEG) polymers to protein molecules to improve various pharmacological properties of the proteins. This modification technique has widespread applications in drug delivery and therapy. Advantages: Increased in vivo half-life: PEGylation can enhance the stability and half-life of proteins in the body, thereby reducing the frequency of administration. Reduced immunogenicity: Due to PEG’s non-immunogenic nature, it can ......

• • How Much Does the Molecular Weight Increase After Protein Glycosylation

  Protein glycosylation is the process of covalently attaching glycans to a protein. This modification increases the molecular weight of the protein, with the exact increment depending on the following factors: 1. Type and Number of Glycans Different types of glycans (such as glucose, galactose, and N-acetylglucosamine) have distinct molecular weights, and multiple glycans can be attached to a single protein. 2. Length and Branching of Glycan Chains Glycan chains may consist of a single monosaccharide........