Related Experiment Video
Updated: Mar 24, 2026
12:19
Tools to Study the Role of Architectural Protein HMGB1 in the Processing of Helix Distorting, Site-specific DNA Interstrand Crosslinks
Published on: November 10, 2016
8.7K
Structural and functional characterization of a hyperthermostable single-stranded DNA-binding protein from a hot
Olesia Werbowy1, Maria Håkansson2, Sebastian Dorawa3
1Department of Microbiology, Faculty of Biology, University of Gdansk, Gdansk, Poland.
Protein Science : a Publication of the Protein Society
|March 23, 2026
Summary
We discovered SSB-M5, a heat-stable single-stranded DNA-binding protein from Iceland. This protein binds DNA with high affinity and improves PCR amplification, showing potential for biotechnology.
Area of Science:
- Biochemistry
- Structural Biology
- Molecular Biology
Background:
- Single-stranded DNA-binding proteins (SSBs) are crucial for DNA replication, repair, and recombination.
- Hyperthermophilic organisms possess unique proteins adapted to extreme temperatures, offering potential biotechnological applications.
Purpose of the Study:
- To structurally and functionally characterize a novel single-stranded DNA-binding protein (SSB-M5) from a hot spring metagenome.
- To assess the DNA-binding properties and thermal stability of SSB-M5.
- To evaluate the utility of SSB-M5 in polymerase chain reaction (PCR) applications.
Main Methods:
- Overproduction and purification of recombinant SSB-M5.
- X-ray crystallography for structural determination.
- Electrophoretic mobility shift assays (EMSA) and quantitative binding analyses.
- Functional complementation assay in Escherichia coli and PCR enhancement assays.
Main Results:
- SSB-M5 forms a functional tetramer and exhibits high thermal stability, remaining active up to 100°C.
- SSB-M5 binds single-stranded DNA with very high affinity (KD = 72 ± 6 pM) and displays cooperative binding.
- SSB-M5 successfully substituted for E. coli SSB in vivo and significantly improved PCR specificity and yield in vitro.
Conclusions:
- SSB-M5 is a hyperthermostable, high-affinity single-stranded DNA-binding protein with a tetrameric functional unit.
- Its robust DNA-binding capabilities and thermal stability make it a promising candidate for molecular biology and biotechnology applications, particularly in PCR.
- This study expands the repertoire of characterized extremophilic DNA-binding proteins.
Related Concept Videos
01:29Diversity of Archaea IV
594
Hyperthermophilic archaea are a group of extremophiles thriving at temperatures above 80°C, often in hydrothermal vents and volcanic soils where conditions surpass the boiling point of water. At such temperatures, proteins, membranes, and DNA in most organisms degrade, but hyperthermophiles have evolved remarkable adaptations to maintain stability and function.Unique Cellular FeaturesHyperthermophilic membranes are composed of a monolayer of biphytanyl tetraether lipids, which resist...
594
01:27Diversity of Archaea III
430
Crenarchaeota, a prominent phylum of Archaea, is remarkable for its ability to thrive in extreme environments characterized by high temperatures and acidity. These microorganisms inhabit sulfuric hot springs, volcanic systems, and submarine hydrothermal vents, where temperatures often exceed 100°C. The unique adaptations of Crenarchaeota not only allow survival under such extreme conditions but also provide insights into the mechanisms of life in primordial Earth-like...
430
01:21Hyperthermophilic Bacteria
718
Domain Bacteria includes some unique hyperthermophilic species. They exhibit remarkable adaptations that enable survival in extreme environments.Thermotoga species are rod-shaped, gram-negative, non-sporulating hyperthermophiles that form a sheath-like envelope called a toga. They ferment sugars or starch, producing lactate, acetate, CO₂, and H₂, and can also grow via anaerobic respiration using H₂ and ferric iron. Found in hot springs and hydrothermal vents, over 20% of their...
718
01:03Single-Strand DNA Binding Proteins
17.1K
For successful DNA replication, the unwinding of double-stranded DNA must be accompanied by stabilization and protection of the separated single strands of the DNA. This crucial task is performed by single-strand DNA-binding (SSB) proteins. They bind to the DNA in a sequence-independent manner, which means that the nitrogenous bases of the DNA need not be present in a specific order for binding of SSB proteins to it. The binding of SSB proteins straightens single-stranded DNA (ssDNA) and makes...
17.1K
01:30Diversity of Archaea I
831
Archaea, a domain of single-celled microorganisms, are classified into five major phyla based on genetic and biochemical characteristics: Euryarchaeota, Crenarchaeota, Thaumarchaeota, Korarchaeota, and Nanoarchaeota. Among these, the phylum Euryarchaeota is notable for its remarkable diversity in morphology, metabolism, and ecological adaptations.Morphological and Metabolic DiversityMembers of Euryarchaeota exhibit a variety of cellular shapes, including rods and cocci. Their metabolic pathways...
831
00:55DNA Helicases
24.8K
DNA unwinding helicase enzymes are a type of motor protein. Motor proteins can translocate along filaments or polymers using energy generated from ATP hydrolysis. Helicases are involved in all the important cellular processes where DNA unwinding is required, such as DNA replication, repair, recombination, and transcription. They are present in all living organisms, but vary in their structure, function, and mechanism of action. For example, in prokaryotes, DnaB helicase binds and translocates...
24.8K