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CRB Discovery
  In this issue
Spectrum™ Peptides:
Cambridge Research Biochemicals offers the most comprehensive range of fluorescent dyes available for peptide labelling
Focus on FRET:
A published CSP-1 FRET reporter peptide synthesised by Cambridge Research Biochemicals
Fluorescent Dye Selector:
Cambridge Research Biochemicals' interactive online dye selector tool

 
Spectrum PeptidesSpectrum Peptides
fluorescent labeling

Cambridge Research Biochemicals offer the worlds’ most comprehensive selection of fluorescent dyes from leading manufacturers including Life Technologies (Invitrogen-Molecular Probes, now Thermo Fisher Scientific), GE Healthcare (Amersham Biosciences), Licor Biosciences, Biosearch Technologies, Cyanagen and Atto-Tec. Both proprietary and non-proprietary dyes are offered, spanning the entire spectrum and technical assistance can be provided in order to select the optimum dye or dye-quencher pair for your application.
Available Proprietary Dyes Include:
  • Life Technologies Dyes: Alexa Fluor®*, Bodipy®*, Oregon Green®
  • GE Healthcare: CyDyes™*
  • Atto-Tec: ATTO dyes
  • Cyanagen: Chromis dyes
  • Biosearch Technologies: CAL Fluor®, Quasar®
  • Licor Biosciences: IR Dyes®*­
* Requires payment of a royalty.

Cambridge Research Biochemicals also offer non-proprietary dyes (no licence fees or royalty payments required). These include:
  • FAM
  • TAMRA
  • EDANS
  • Mca
  • FITC
Click Chemistry
Cambridge Research Biochemicals have recently signed a non-exclusive license with The Scripps Research Institute (TSRI) to add click chemistry services, namely the “click” labelling of peptides and proteins, to its existing service portfolio.This technology traditionally involves coupling of an alkyne functionality within an organic system to an azide moiety using a copper catalyst, although ‘copper-free’ click chemistry is now widely used.

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Technical Excellence:
  • 0.5mg to multi-mg scale
  • Dye attachment to the N-terminus, C-terminus or side-chain of the peptide using amino-, thiol-, or click-reactive derivatives
  • Optimised strategy for synthesis of FRET peptides to minimise background fluorescence
  • ISO9001:2008 certified
  • Rapid turnaround – typically within 3-4 weeks
  • Please visit our Technical pages for more information

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  Focus on Fret  
Using a CSP-1 (Competence-Stimulating Peptide)  FRET reporter peptide synthesised by
Cambridge Research Biochemicals Sebert et al at the Children’s Hospital of Philadelphia have determined that the presence of proteins with exposed hydrophobic regions competitively reduces the degradation of competence-stimulating peptide (CSP). This suggests that a mechanism by which The HtrA protease of Streptococcus pneumoniae functions as a sensor for biosynthetic errors is taking place, as HtrA digests both the CSP and unfolded proteins.

To generate the peptide,
[QSY-7]-EMRLSKFFRDFILQRKK-[Cys(Alexa 488)]-amide, Cambridge Research Biochemicals functionalised CSP-1 with a QSY-7 quencher (Life Technologies) at the N-terminus and an Alexa 488 (Life Technologies) fluorophore via cysteine at the C terminus. The QSY-7 NHS ester quencher was manually coupled on-resin to the N-terminus and the intermediate peptide cleaved and purified prior to addition of the AF488 maleimide dye in solution. To achieve good sensitivity, FRET peptides must be at high purity and free from residual dye, so the target FRET peptide was further purified to >95% purity by HPLC following dye-conjugation and its identity verified by mass spectrometry.
 
Diagram of A FRET substrate using a fluorophore as a donor and non-fluorescent quencher.
Diagram: A FRET substrate using a fluorophore as a donor and a non-fluorescent (dark) quencher as an acceptor. Enzymatic cleavage results in an increase in donor signal - Sebert et al proved by mass spectrometry that cleavage predominantly followed residue Phe-8 of the CSP-1 isoform of the peptide within its central hydrophobic patch.

Reference:
Pneumococcal HtrA Digests Denatured Proteins and CSP
Cassone M, Gagne AL, Spruce LA, Seeholzer SH, Sebert ME
Journal of Biological Chemistry, 2012, 287(46): 38449-38459
Fluorescent Dye Selector
Cambridge Research Biochemicals' interactive online dye selector wheel will guide you through the process of selecting the optimum dye for your application.
Try our online fluorescent dye selector wheel at http://www.crbdiscovery.com/services/dye-labelling/ and select your desired wavelength to reveal Cambridge Research Biochemicals' comprehensive selection of dyes at each region of the spectrum.

Cambridge Research Biochemicals offer dyes compatible with all commonly used excitation sources. Those listed are the most commonly requested, however do get in touch if your preferred dye is not listed.
 
  Fluorescent Dye Selector

We offer dyes compatible with all commonly used excitation sources. The most frequently requested are listed, together with the appropriate quencher for FRET applications. If you require a peptide labelled with a dye which is not listed, please enquire, as we have access to many more.
To order your spectrum peptide please contact us via email, telephone or visit our website
CRB will be attending and exhibiting at the following conferences and seminars:
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Request a Quotation
Should you require a quotation for any of our custom services, we have a clear cut and easy system that enables you to do so.
 
At Cambridge Research Biochemicals we have a fast-track quotation system that tailors your requirements, gives you all the information you need and a specific price within 24 hours. You are also given a personal contact so that if you have any further questions you can directly contact one of our approachable representatives who has the technical knowledge to assist you.
1   Go to: www.crbdiscovery.com/quotations
     
2   Click on the relevant tab and register your details.
     
3   Once issued with a password log on to our quotation system using your customer account log-in and a quotation will be issued within 24 hours.