A novel chimeric recombinant protein PDHB-P80 of Mycoplasma agalactiae as a potential diagnostic tool

نویسندگان

  • Farhid Hemmatzadeh School of Animal and Veterinary Sciences, The University of Adelaide, SA, Australia
  • Malihe Akbarzadeh-Niaki Department of Pathobiology, Biotechnology Section, School of Veterinary Medicine, Shiraz University, Shiraz, Iran
  • Nasrin Kazemipour Department of Basic Sciences, School of Veterinary Medicine, Shiraz University, Shiraz, Iran
  • Vida Eraghi Department of Pathobiology, Biotechnology Section, School of Veterinary Medicine, Shiraz University, Shiraz, Iran
چکیده مقاله:

The aim of this study was to construct, expression of a novel recombinant chimeric protein consisting of Pyruvate dehydrogenase beta subunit (PDHB) and high antigenic region of integral membrane lipoprotein P80 of Mycoplasma agalactiae as a potential diagnostic tool. The full-length sequence of pdhb and a portion of antigenic regions of P80 were selected and analyzed by CLC main workbench 5.5 software. Several linkers and three dimensional structure of PDHB-P80 were compared to the native PDHB and analyzed to select a proper one for expression. The fusion gene sequence was optimized and synthesized in pMAT cloning vector. The synthetic pMAT-pdhb-p80 was digested using Bam HI and Sal I restriction enzymes and ligated into pMAL-p5X expression vector. The pMAL-pdhb-p80 construct was transfected into E.coli BL21 strain cells and expressed protein were purified using amylose resin. and the purified protein was analyzed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. In silico analysis demonstrated that fusion proteins using IgG4 middle hinge (CPSCP) with TM-score of 0.99 showed the higher similarity between three dimensional structure of PDHB before and after fusion with high antigenic region of P80. Successful cloning verified by PCR colony, double digestion and sequence analysis. Besides, SDS-PAGE analysis and Western blotting indicated and confirmed the expression of intact recombinant chimeric protein MBP-PDHB-P80 along with some truncated forms of the recombinant protein. it could be concluded that the fusion construct has a potential for serodiagnostic assay in future studies.

برای دانلود باید عضویت طلایی داشته باشید

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

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Expression of a New Recombinant Collagenase Protein of Lucilia Sericata in SF9 Insect Cell as a Potential for Wound Healing

Background: Today, the use of maggot therapy has become widespread due to the increase in chronic ulcers in the world. The recombinant production of secreted enzymes from these larvae is a novel non-invasive method for the treatment of chronic ulcers. Lucilia sericata (L. sericata) collagenase (MMP-1) has been expressed in insect cells. Collagenase is an enzyme that is widely used in clinical t...

متن کامل

expression and purification of the recombinant cytochrome p450 cyp141 protein of mycobacterium tuberculosis as a diagnostic tool and vaccine production

conclusions in this experimental study, for the first time in iran the expression and purification of this recombinant protein was done successfully. this recombinant protein could be used as a vaccine candidate and diagnostic purpose in subsequent investigations. results the highest expression of the cytochrome p450 cyp141 protein was obtained by the addition of 1 mm of isopropyl β-d-1-thiogal...

متن کامل

Design and Production of a Novel Recombinant Chimeric IL2-Omp31 Antigen against Brucella Infection

Brucellosis is a zoonotic disease in human and animals. Brucella melitensis is one of the most pathogenic species of Brucella in goat and sheep. Omp31 is an outer membrane protein of Brucella that acts as an immunogenic protein. Cytokines are glycoproteins with low molecular weight that play the role of an immune adjuvant and regulate immune responses. Interleukin-2 is one of the most important...

متن کامل

characterization of afb, a novel bifunctional protein in streptococcus agalactiae

background and objectives: streptococcus agalactiae is the leading cause of bacterial sepsis and meningitis in newborns and results in pneumonia and bacteremia in adults. a number of s. agalactiae components are involved in colonization of target cells. destruction of peptidoglycan and division of covalently linked daughter cells is mediated by autolysins. in this study, autolytic activity and ...

متن کامل

Saliva As a Diagnostic Tool in Covid 19 Detection, A Review

Background and Aim: Delayed detection of SARS-CoV-2 has been a major hindrance in curbing the current COVID-19 pandemic. Wide spread testing is the most practical way out of the current crisis. The current gold standard for COVID-19 diagnosis is real-time reverse transcription polymerase chain reaction (RT-PCR) detection of SARS-CoV-2 from nasopharyngeal swabs. However, collecting nasopharyngea...

متن کامل

wuthering heights and the concept of marality/a sociological study of the novel

to discuss my point, i have collected quite a number of articles, anthologies, and books about "wuthering heights" applying various ideas and theories to this fantastic story. hence, i have come to believe that gadamer and jauss are rightful when they claim that "the individaul human mind is the center and origin of all meaning," 3 that reading literature is a reader-oriented activity, that it ...

15 صفحه اول

منابع من

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

ذخیره در منابع من قبلا به منابع من ذحیره شده

{@ msg_add @}


عنوان ژورنال

دوره 9  شماره 3

صفحات  123- 128

تاریخ انتشار 2020-09-01

با دنبال کردن یک ژورنال هنگامی که شماره جدید این ژورنال منتشر می شود به شما از طریق ایمیل اطلاع داده می شود.

میزبانی شده توسط پلتفرم ابری doprax.com

copyright © 2015-2023