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Search for a dextransucrase minimal motif involved in dextran binding
Surisa Suwannarangsee1, Claire Moulis, Gabrielle Potocki-Veronese
1Biological Science Program, Faculty of Science, Chulalongkorn University, 254 Phyathai Road, Patumwan, Bangkok, Thailand.
FEBS Letters
|September 11, 2007
Summary
Researchers identified a small dextransucrase fragment (GBD-7) that strongly binds dextran polymers. This finding supports the role of the GBD domain in dextran formation and suggests GBD-7
Area of Science:
- Enzymology
- Biochemistry
- Molecular Biology
Background:
- Dextransucrase from Leuconostoc mesenteroides NRRL B512-F is crucial for dextran synthesis.
- Understanding the dextran-binding properties of dextransucrase is key to elucidating its function in polymer formation.
- Truncated enzyme forms can help pinpoint functional domains within larger proteins.
Purpose of the Study:
- To investigate the dextran-binding capabilities of various truncated dextransucrase forms.
- To identify the minimal sequence responsible for strong dextran interaction.
- To evaluate the potential of identified fragments for biotechnological applications, such as affinity purification.
Main Methods:
- Construction and expression of fourteen truncated variants of Leuconostoc mesenteroides NRRL B512-F dextransucrase.
- Assessment of dextran-binding activity for each truncated form.
- Determination of dissociation constants (K(d)) for high-affinity fragments using techniques like surface plasmon resonance (SPR) or isothermal titration calorimetry (ITC).
Main Results:
- A 14kDa fragment, GBD-7 (amino acids N1397-A1527), demonstrated strong dextran-binding affinity.
- GBD-7 exhibits a high affinity for isomaltohexaose and longer dextran chains, with a K(d) of 2.8x10(-9)M.
- The six YG repeats within GBD-7 are crucial for this high-affinity interaction.
Conclusions:
- The C-terminal dextran-binding domain (GBD) plays a significant role in dextran polymer formation.
- GBD-7 represents the smallest functional unit for strong dextran binding, confirming its importance.
- GBD-7 shows potential as an affinity tag for efficient dextran purification using inexpensive resins.
