Analysis Biomolecular Interactions of PirA and PII from Synechocystis by BLI (CAT#: STEM-MB-0176-CJ)

Introduction

Synechocystis is a genus of unicellular, freshwater cyanobacteria in the family Merismopediaceae. Like all cyanobacteria, Synechocystis branches on the evolutionary tree from its ancestral root, Gloeobacter violaceus. Synechocystis is not diazotrophic, and is closely related to another model organism, Cyanothece 51442. In addition, the accumulation of Ssr0692, a small protein in Synechocystis, is a response to ammonium supply and plays a key regulatory role in cyanobacterial metabolism through its interaction with PII signalling proteins. It can directly interfere with NAGK-dependent activation of pii. Consistent with this, at fluctuating nitrogen levels, the ssr0692 mutant strain is impaired in balancing the synthesis of arginine and other amino acids that are associated with the cyanobacterial ornithine ammonia cycle.

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Principle Applications Procedure Materials Notes Related Products

Principle

Bio-Layer Interferometry (BLI) is an optical technique for measuring macromolecular interactions by analyzing interference patterns of white light reflected from the surface of a biosensor tip. BLI experiments are used to determine the kinetics and affinity of molecular interactions. In a BLI experiment, one molecule is immobilized to a Dip and Read Biosensor and binding to a second molecule is measured. A change in the number of molecules bound to the end of the biosensor tip causes a shift in the interference pattern that is measured in real-time.

Applications

Botany

Procedure

1. Detect Buffers and prepare samples. BLI experiments are set up with one molecule immobilised on the surface of the biosensor (load sample) and a second molecule in solution (the analytical sample).
2. Fix the load sample on the biocompatible biosensor while the analytical sample is in solution.
3. The biosensor tip is immersed in the solution so that the target molecule begins to bind to the analysis sample.
4. Set up and run the BLI experiment. Molecules bound to or dissociated from the biosensor can generate response curves on the BLI system; unbound molecules, changes in the refractive index of the surrounding medium or changes in flow rate do not affect the interferogram pattern.
5. Collect and analyse data on the BLI's system.

Materials

• Equipment: Gator® Bio-Layer Interferometry (BLI)
• Sample Type: DNA, RNA, Protein, Antibodies, Peptides, Small Molecules

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