English (United Kingdom)

https://curated-unify.zendy.io/wp-json/zendy-region/v1/featured_content/oa?rat=en

https://curated-unify.zendy.io/wp-json/zendy-region/v1/highlighted_journal/

Global: Zendy Plus annual

Presents the access of premium content as premium feature

Premium Content

Presents the keyphrase highlighting as premium feature

Keyphrase Highlighting

Presents the summarisation as premium feature

Summarisation

Insights

Presents the pdf analysis as premium feature

PDF Analysis

Presents the zaia usage as premium feature

ZAIA

Global: Zendy Tools annual

Global: Zendy Free Plan

Global: Zendy Tools monthly

Global: Zendy Plus monthly

Background  Biotin is an essential enzyme cofactor that acts as a CO 2  carrier in carboxylation and decarboxylation reactions. The  E. coli  genome encodes a biosynthetic pathway that produces biotin from pimeloyl-CoA in four enzymatic steps. The final step, insertion of sulfur into desthiobiotin to form biotin, is catalyzed by the biotin synthase, BioB. A dedicated biotin ligase (BirA) catalyzes the covalent attachment of biotin to biotin-dependent enzymes. Isotopic labeling has been a valuable tool for probing the details of the biosynthetic process and assaying the activity of biotin-dependent enzymes, however there is currently no established method for  35 S labeling of biotin.    Results  In this study, we produced [ 35 S]-biotin from Na 35 SO 4  and desthiobiotin with a specific activity of 30.7 Ci/mmol, two orders of magnitude higher than previously published methods. The biotinylation domain ( Pf BCCP-79) from the  Plasmodium falciparum  acetyl-CoA carboxylase (ACC) was expressed in  E. coli  as a biotinylation substrate. We found that overexpression of the  E. coli  biotin synthase, BioB, and biotin ligase, BirA, increased  Pf BCCP-79 biotinylation 160-fold over basal levels. Biotinylated  Pf BCCP-79 was purified by affinity chromatography, and free biotin was liberated using acid hydrolysis. We verified that we had produced radiolabeled biologically active [ D ]-biotin that specifically labels biotinylated proteins through reuptake in  E. coli .    Conclusions  The strategy described in our report provides a simple and effective method for the production of [ 35 S]-biotin in  E. coli  based on affinity chromatography.

Overexpression of biotin synthase and biotin ligase is required for efficient generation of sulfur-35 labeled biotin in E. coli