Identification of the critical residues responsible for differential reactivation of the triosephosphate isomerases of two trypanosomes

نویسندگان

  • Monica Rodríguez-Bolaños
  • Nallely Cabrera
  • Ruy Perez-Montfort
چکیده

The reactivation of triosephosphate isomerase (TIM) from unfolded monomers induced by guanidine hydrochloride involves different amino acids of its sequence in different stages of protein refolding. We describe a systematic mutagenesis method to find critical residues for certain physico-chemical properties of a protein. The two similar TIMs of Trypanosoma brucei and Trypanosoma cruzi have different reactivation velocities and efficiencies. We used a small number of chimeric enzymes, additive mutants and planned site-directed mutants to produce an enzyme from T. brucei with 13 mutations in its sequence, which reactivates fast and efficiently like wild-type (WT) TIM from T. cruzi, and another enzyme from T. cruzi, with 13 slightly altered mutations, which reactivated slowly and inefficiently like the WT TIM of T. brucei Our method is a shorter alternative to random mutagenesis, saturation mutagenesis or directed evolution to find multiple amino acids critical for certain properties of proteins.

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Correction to ‘Identification of the critical residues responsible for differential reactivation of the triosephosphate isomerases of two trypanosomes’

to 'Identification of the critical residues responsible for differential reactivation of the triosephosphate isomerases of two trypanosomes'. (c) mutant no. additive mutations in region 2 regions of TcTIM scheme of different amino acids in region 2

متن کامل

Three unrelated and unexpected amino acids determine the susceptibility of the interface cysteine to a sulfhydryl reagent in the triosephosphate isomerases of two trypanosomes

Proteins with great sequence similarity usually have similar structure, function and other physicochemical properties. But in many cases, one or more of the physicochemical or functional characteristics differ, sometimes very considerably, among these homologous proteins. To better understand how critical amino acids determine quantitative properties of function in proteins, the responsible res...

متن کامل

Identification of Amino Acids that Account for Long-Range Interactions in Two Triosephosphate Isomerases from Pathogenic Trypanosomes

For a better comprehension of the structure-function relationship in proteins it is necessary to identify the amino acids that are relevant for measurable protein functions. Because of the numerous contacts that amino acids establish within proteins and the cooperative nature of their interactions, it is difficult to achieve this goal. Thus, the study of protein-ligand interactions is usually f...

متن کامل

REASSOCIATION AND REACTIVATION OF GLUCOSE 6-PHOSPHATE DEHYDROGENASE FROM STREPTOMYCES AUREOFACIENS AFTER DENATURATION BY 6 M UREA

Glucose 6-phosphate dehydrogenase (G6PD) from Streptomyces aureofaciens was purified and denatured in 6 M urea. Denaturation led to complete dissociation of the enzyme into its inactive monomers, 98% loss of the enzyme activity, about 30% decrease in the protein fluorescence and a 10 nm red shift in the emission maximum. Dilution of urea-denatured enzyme resulted in regaining of the enzyme acti...

متن کامل

Impact of Cooking Procedures on Antibacterial Drug Residues in Foods: A Review

Antibacterial drugs used in animal might result in deposition of residues in meat, milk and eggs. The presence of antimicrobial residues in animal-originated food is critical problem in many countries over the years. Because, drug residues might result in various health hazards, both actual incidence of reactions and potential hazards perceived by the public, the most countries assess residue o...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

عنوان ژورنال:

دوره 6  شماره 

صفحات  -

تاریخ انتشار 2016