Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

Tagging and Fusion Proteins01:24

Tagging and Fusion Proteins

6.6K
Proteins are involved in several cellular processes and biochemical reactions. Analyzing a specific protein of interest requires it to be isolated from the other proteins in the cell. This is achieved by overexpressing the specific gene in a suitable host to produce large quantities of the target protein. A tag or label is recombined with the gene to produce a fusion protein containing the target protein and the tag. The tags on these fusion proteins can then be used for easy detection and...
6.6K
Tail-anchoring of Proteins in the ER Membrane01:45

Tail-anchoring of Proteins in the ER Membrane

3.1K
Tail-anchored, or TA, proteins are estimated to make up to 3-5% of membrane proteins found in the eukaryotic cell. Such proteins have a single transmembrane domain located approximately 30 amino acid residues upstream from the C-terminal end. As a result, the signal recognition particle (SRP) cannot guide a TA protein to the ER membrane for cotranslational insertion. Hence, they are integrated into the ER membrane post-translationally using their C-terminal end as the anchor. TA proteins...
3.1K
From DNA to Protein03:06

From DNA to Protein

18.4K
The flow of genetic information in cells from DNA to mRNA to protein is described by the central dogma, which states that genes specify the sequence of mRNAs, which in turn specify the sequence of amino acids making up all proteins. The decoding of one molecule to another is performed by specific proteins and RNAs. Because the information stored in DNA is so central to cellular function, it makes intuitive sense that the cell would make mRNA copies of this information for protein synthesis...
18.4K
Leaky Scanning02:28

Leaky Scanning

5.1K
During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA.  Marilyn Kozak discovered that the sequence RCCAUGG (where R...
5.1K
Transfer RNA Synthesis02:36

Transfer RNA Synthesis

11.9K
One of the unique features of tRNA is the presence of modified bases. In some tRNAs, modified bases account for nearly 20% of the total bases in the molecule. Altogether, these unusual bases protect the tRNA from enzymatic degradation by RNases.
Each of these chemical modifications is carried by a specific enzyme, post-transcription. All of these enzymes have unique base and site-specificity. Methylation, the most common chemical modification, is carried by at least nine different enzymes, with...
11.9K
Translocation of Proteins into the Mitochondria01:19

Translocation of Proteins into the Mitochondria

3.1K
Mitochondrial precursors are translocated to the internal subcompartments via independent mechanisms involving distinct protein machineries called translocases.
Sorting of outer membrane proteins:
Mitochondrial outer membrane proteins are of two types: the transmembrane, beta-barrel porins, and the membrane-anchored, alpha-helical proteins. Beta-barrel porin precursors are translocated by the TOM complex and inserted into the outer mitochondrial membrane by the SAM complex. In contrast,...
3.1K

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Tracking Down the Evolution of Microorganisms by Exhaustive Bottom-Up Analysis of Proteomes.

International journal of molecular sciences·2026
Same author

Multidimensional Gene Space as an Approach for Analyzing the Organization of Genomes.

International journal of molecular sciences·2025
Same author

Engineering of a Complex of the DNase Domain of Colicin E9 with the Immunity Protein Im9 Activated by the Protease pS273R of African Swine Fever Virus.

ACS synthetic biology·2025
Same author

Discovery of Bivalent GalNAc-Conjugated Betulin as a Potent ASGPR-Directed Agent against Hepatocellular Carcinoma.

Bioconjugate chemistry·2021
Same author

Mannosylated Cationic Copolymers for Gene Delivery to Macrophages.

Macromolecular bioscience·2021
Same author

New Small-Molecule Glycoconjugates of Docetaxel and GalNAc for Targeted Delivery to Hepatocellular Carcinoma.

Molecular pharmaceutics·2020

Related Experiment Video

Updated: Jul 5, 2025

A Fast and Quantitative Method for Post-translational Modification and Variant Enabled Mapping of Peptides to Genomes
09:10

A Fast and Quantitative Method for Post-translational Modification and Variant Enabled Mapping of Peptides to Genomes

Published on: May 22, 2018

9.2K

Novel His-tag Variants for Insertion Inside Polypeptide Chain.

Anastasiia G Tarabarova1, Anton Lopukhov2, Alexey N Fedorov3

  • 1A N Bach Institute of Biochemistry of the Russian Academy of Sciences, Leninskii prosp 33/2, Moscow 119071, Russian Federation.

ACS Omega
|January 15, 2024
PubMed
Summary

Newly designed internal His-tags, featuring histidine and glycine residues, minimize protein structural impact. These insertable tags offer a new method for purifying diverse proteins without N- or C-terminal fusion.

More Related Videos

Residue-specific Incorporation of Noncanonical Amino Acids into Model Proteins Using an Escherichia coli Cell-free Transcription-translation System
11:47

Residue-specific Incorporation of Noncanonical Amino Acids into Model Proteins Using an Escherichia coli Cell-free Transcription-translation System

Published on: August 1, 2016

16.0K
Residue-Specific Exchange of Proline by Proline Analogs in Fluorescent Proteins: How "Molecular Surgery" of the Backbone Affects Folding and Stability
10:31

Residue-Specific Exchange of Proline by Proline Analogs in Fluorescent Proteins: How "Molecular Surgery" of the Backbone Affects Folding and Stability

Published on: February 3, 2022

3.0K

Related Experiment Videos

Last Updated: Jul 5, 2025

A Fast and Quantitative Method for Post-translational Modification and Variant Enabled Mapping of Peptides to Genomes
09:10

A Fast and Quantitative Method for Post-translational Modification and Variant Enabled Mapping of Peptides to Genomes

Published on: May 22, 2018

9.2K
Residue-specific Incorporation of Noncanonical Amino Acids into Model Proteins Using an Escherichia coli Cell-free Transcription-translation System
11:47

Residue-specific Incorporation of Noncanonical Amino Acids into Model Proteins Using an Escherichia coli Cell-free Transcription-translation System

Published on: August 1, 2016

16.0K
Residue-Specific Exchange of Proline by Proline Analogs in Fluorescent Proteins: How "Molecular Surgery" of the Backbone Affects Folding and Stability
10:31

Residue-Specific Exchange of Proline by Proline Analogs in Fluorescent Proteins: How "Molecular Surgery" of the Backbone Affects Folding and Stability

Published on: February 3, 2022

3.0K

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein Engineering

Background:

  • Hexahistidine (His-tag) affinity purification is a standard technique.
  • N- or C-terminal His-tag fusion can negatively affect protein structure and function.
  • Internal tag insertion is explored to mitigate N/C-terminal fusion issues.

Purpose of the Study:

  • To design and characterize novel internal His-tags.
  • To minimize steric hindrance and impact on protein structure/function.
  • To broaden the utility of fusion proteins like eGFP and GrAD207.

Main Methods:

  • Designed internal His-tags with two histidine triplets separated by glycine residues.
  • Tested tag applicability by fusing them internally into enhanced green fluorescent protein (eGFP).
  • Validated tag functionality with a modified GroEL chaperone domain (GrAD207).

Main Results:

  • Successfully incorporated internal His-tags into eGFP and GrAD207.
  • Demonstrated that internal tags minimize steric hindrance compared to terminal tags.
  • Showed the potential for broad applicability across different protein classes.

Conclusions:

  • Internal His-tags offer a viable alternative to N/C-terminal fusions.
  • The designed tags can be adjusted for efficient purification of diverse proteins.
  • This approach broadens the functionality of fusion proteins without compromising termini.