CONFOCHECK - Dedicated Fourier Transform Infrared (FT-IR) Spectrometer Developed for Protein Analysis in Aqueous Solution by Bruker Optics

Topics Covered

About Bruker Optics
Infrared Protein Analysis
Advantages of Infrared Protein Analysis
Applications

About Bruker Optics

Bruker Optics, part of the Bruker Corporation (NASDAQ:BRKR) is the leading manufacturer and worldwide supplier of Fourier Transform Infrared, Near Infrared and Raman spectrometers for various industries and applications.

Bruker entered the field of FT-IR spectroscopy in 1974. The early instruments set new standards in research FT-IR with evacuable optics, high resolution and automatic range change. Since then, the product line has been continuously expanding with instruments suitable for both analytical and research applications with exceptional performance characteristics.

Today, Bruker Optics offers FT-IR, NIR, Raman, TD-NMR, TeraHertz spectrometers and imaging spectrographs for various markets and applications. Bruker Optics has R&D and manufacturing centers in Ettlingen, Germany and The Woodlands, USA, technical support centers and sales offices throughout Europe, North and South America and Asia.

Infrared Protein Analysis

In infrared spectroscopy molecular vibrations are measured due to the specific absorption of infrared radiation by chemical bonds. The shape and frequency of the amide I band, which is assigned to the C=O stretching vibration within the peptide bonds of the protein molecule, is very characteristic for the structure of the studied protein. From this single band the presence and relative percentage of secondary structure elements (alpha-helix, beta-sheet) in a protein can be determined and conformational changes are detected with high sensitivity. Any alterations in the structure regardless if they were induced by temperature or pH changes, ligand binding, mutations etc. are reflected by specific absorptive changes in the amide I region.

Since IR obeys the rules of Lambert-Beer´s law, the protein spectrum can be utilized for concentration analysis. Within seconds and without any bio- or immuno-chemical staining, protein concentrations in an extremely wide range from about 0.1 mg/ml to >100 mg/ml can be determined directly from the undiluted solution.

However, the information content of IR protein spectrum is not limited to the amide I band. Side chains of various amino acids can also be monitored and directly compared under varying environmental conditions (pH, salts, ligands), or in mutant proteins with altered primary sequence.

Advantages of Infrared Protein Analysis

The advantages of infrared protein analysis are its ease of use and its relatively inexpensive capability of studying proteins in aqueous media at any buffer conditions. Therefore, the IR-technique is a powerful analysis method in the pharmaceutical industry, e.g. for formulation optimization, stability studies during drug development and QC of protein drug products.

In research isolate structural effects of a protein while binding to drugs or interacting with substrates and inhibitors are followed. Furthermore, the formation of multimeric structures like aggregates or fibrils can be monitored by infrared spectroscopy (IR), even in real time.

It should be stressed that these investigations are not restricted to water-soluble proteins, but can also be performed on membrane bound proteins.

The CONFOCHECK is a dedicated Fourier transform infrared (FT-IR) spectrometer system that was developed to fulfill the demanding requirements for protein analysis in aqueous solution. Its specific concept facilitates a fast data acquisition with a high sample throughput controlled by a user friendly software interface.

Applications

Applications for CONFOCHECK are:

  • Protein dynamics (temperature induced conformational changes)
  • Protein quantification (from 0.03 mg/ml up to > 100mg/ml in aqueous solution)
  • Determination of secondary structure
  • Monitoring of conformational changes during protein aggregation, precipitation and crystallization
  • Quantification of all kind of solutes in aqueous samples

Source Bruker Optics

For more information on this source please visit Bruker Optics

Date Added: May 11, 2010 | Updated: Sep 20, 2013
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