Fusion of selected regions of mycobacterial antigens for enhancing sensitivity in serodiagnosis of tuberculosis

Madeeha Afzal, Sana Khurshid, Ruqyya Khalid, Rehan Zafar Paracha, Imran Khan, M. Waheed Akhtar

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Serodiagnosis of tuberculosis requires detection of antibodies against multiple antigens of Mycobacterium tuberculosis, because antibody profiles differ among the patients. Using fusion proteins with epitopes from two or more antigens would facilitate in the detection of multiple antibodies. Fusion constructs tn1FbpC1-tnPstS1 and tn2FbpC1-tnPstS1 were produced by linking truncated regions of variable lengths from FbpC1 to the N-terminus of the truncated PstS1. Similarly a truncated fragment of HSP was linked to the N-terminus of a truncated fragment from FbpC1 to produce tnHSP-tn1FbpC1. ELISA analysis of the plasma samples of TB patients against tn2FbpC1-tnPstS1 showed 72.2% sensitivity which is nearly the same as the expected combined value for the two individual antigens. However, the sensitivity of tn1FbpC1-tnPstS1 was lowered to 60%. tnHSP-tn1FbpC1 showed 67.7% sensitivity which is slightly less than the expected combined value for the two individual antigens, but still significantly higher than that of each of the individual antigen. Data for secondary structure analysis by CD spectrometry was in reasonable agreement with the X-ray crystallographic data of the native proteins and the predicted structure of the fusion proteins. Comparative molecular modeling suggests that the epitopes of the constituent proteins are better exposed in tn2FbpC1-tnPstS1 as compared to those in tn1FbpC1-tnPstS1. Therefore, removal of the N-terminal non-epitopic region of FbpC1 from 34-96 amino acids seems to have unmasked at least some of the epitopes, resulting in greater sensitivity. The high level of sensitivity of tn2FbpC1-tnPstS1 and tnHSP-tn1FbpC1, not reported before, shows that these fusion proteins have great potential for use in serodiagnosis of tuberculosis.

Original languageEnglish (US)
Pages (from-to)104-111
Number of pages8
JournalJournal of Microbiological Methods
Volume115
DOIs
StatePublished - Aug 1 2015

Fingerprint

Serologic Tests
Tuberculosis
Antigens
Epitopes
Proteins
Antibodies
Spectrum Analysis
Enzyme-Linked Immunosorbent Assay
X-Rays
Amino Acids

Keywords

  • Epitopes
  • FbpC1
  • Fusion antigens
  • HSP
  • PstS1
  • Serodiagnosis

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology
  • Microbiology (medical)

Cite this

Fusion of selected regions of mycobacterial antigens for enhancing sensitivity in serodiagnosis of tuberculosis. / Afzal, Madeeha; Khurshid, Sana; Khalid, Ruqyya; Paracha, Rehan Zafar; Khan, Imran; Akhtar, M. Waheed.

In: Journal of Microbiological Methods, Vol. 115, 01.08.2015, p. 104-111.

Research output: Contribution to journalArticle

Afzal, Madeeha ; Khurshid, Sana ; Khalid, Ruqyya ; Paracha, Rehan Zafar ; Khan, Imran ; Akhtar, M. Waheed. / Fusion of selected regions of mycobacterial antigens for enhancing sensitivity in serodiagnosis of tuberculosis. In: Journal of Microbiological Methods. 2015 ; Vol. 115. pp. 104-111.
@article{386f7ca747084567b93757cbf29caa5e,
title = "Fusion of selected regions of mycobacterial antigens for enhancing sensitivity in serodiagnosis of tuberculosis",
abstract = "Serodiagnosis of tuberculosis requires detection of antibodies against multiple antigens of Mycobacterium tuberculosis, because antibody profiles differ among the patients. Using fusion proteins with epitopes from two or more antigens would facilitate in the detection of multiple antibodies. Fusion constructs tn1FbpC1-tnPstS1 and tn2FbpC1-tnPstS1 were produced by linking truncated regions of variable lengths from FbpC1 to the N-terminus of the truncated PstS1. Similarly a truncated fragment of HSP was linked to the N-terminus of a truncated fragment from FbpC1 to produce tnHSP-tn1FbpC1. ELISA analysis of the plasma samples of TB patients against tn2FbpC1-tnPstS1 showed 72.2{\%} sensitivity which is nearly the same as the expected combined value for the two individual antigens. However, the sensitivity of tn1FbpC1-tnPstS1 was lowered to 60{\%}. tnHSP-tn1FbpC1 showed 67.7{\%} sensitivity which is slightly less than the expected combined value for the two individual antigens, but still significantly higher than that of each of the individual antigen. Data for secondary structure analysis by CD spectrometry was in reasonable agreement with the X-ray crystallographic data of the native proteins and the predicted structure of the fusion proteins. Comparative molecular modeling suggests that the epitopes of the constituent proteins are better exposed in tn2FbpC1-tnPstS1 as compared to those in tn1FbpC1-tnPstS1. Therefore, removal of the N-terminal non-epitopic region of FbpC1 from 34-96 amino acids seems to have unmasked at least some of the epitopes, resulting in greater sensitivity. The high level of sensitivity of tn2FbpC1-tnPstS1 and tnHSP-tn1FbpC1, not reported before, shows that these fusion proteins have great potential for use in serodiagnosis of tuberculosis.",
keywords = "Epitopes, FbpC1, Fusion antigens, HSP, PstS1, Serodiagnosis",
author = "Madeeha Afzal and Sana Khurshid and Ruqyya Khalid and Paracha, {Rehan Zafar} and Imran Khan and Akhtar, {M. Waheed}",
year = "2015",
month = "8",
day = "1",
doi = "10.1016/j.mimet.2015.06.003",
language = "English (US)",
volume = "115",
pages = "104--111",
journal = "Journal of Microbiological Methods",
issn = "0167-7012",
publisher = "Elsevier",

}

TY - JOUR

T1 - Fusion of selected regions of mycobacterial antigens for enhancing sensitivity in serodiagnosis of tuberculosis

AU - Afzal, Madeeha

AU - Khurshid, Sana

AU - Khalid, Ruqyya

AU - Paracha, Rehan Zafar

AU - Khan, Imran

AU - Akhtar, M. Waheed

PY - 2015/8/1

Y1 - 2015/8/1

N2 - Serodiagnosis of tuberculosis requires detection of antibodies against multiple antigens of Mycobacterium tuberculosis, because antibody profiles differ among the patients. Using fusion proteins with epitopes from two or more antigens would facilitate in the detection of multiple antibodies. Fusion constructs tn1FbpC1-tnPstS1 and tn2FbpC1-tnPstS1 were produced by linking truncated regions of variable lengths from FbpC1 to the N-terminus of the truncated PstS1. Similarly a truncated fragment of HSP was linked to the N-terminus of a truncated fragment from FbpC1 to produce tnHSP-tn1FbpC1. ELISA analysis of the plasma samples of TB patients against tn2FbpC1-tnPstS1 showed 72.2% sensitivity which is nearly the same as the expected combined value for the two individual antigens. However, the sensitivity of tn1FbpC1-tnPstS1 was lowered to 60%. tnHSP-tn1FbpC1 showed 67.7% sensitivity which is slightly less than the expected combined value for the two individual antigens, but still significantly higher than that of each of the individual antigen. Data for secondary structure analysis by CD spectrometry was in reasonable agreement with the X-ray crystallographic data of the native proteins and the predicted structure of the fusion proteins. Comparative molecular modeling suggests that the epitopes of the constituent proteins are better exposed in tn2FbpC1-tnPstS1 as compared to those in tn1FbpC1-tnPstS1. Therefore, removal of the N-terminal non-epitopic region of FbpC1 from 34-96 amino acids seems to have unmasked at least some of the epitopes, resulting in greater sensitivity. The high level of sensitivity of tn2FbpC1-tnPstS1 and tnHSP-tn1FbpC1, not reported before, shows that these fusion proteins have great potential for use in serodiagnosis of tuberculosis.

AB - Serodiagnosis of tuberculosis requires detection of antibodies against multiple antigens of Mycobacterium tuberculosis, because antibody profiles differ among the patients. Using fusion proteins with epitopes from two or more antigens would facilitate in the detection of multiple antibodies. Fusion constructs tn1FbpC1-tnPstS1 and tn2FbpC1-tnPstS1 were produced by linking truncated regions of variable lengths from FbpC1 to the N-terminus of the truncated PstS1. Similarly a truncated fragment of HSP was linked to the N-terminus of a truncated fragment from FbpC1 to produce tnHSP-tn1FbpC1. ELISA analysis of the plasma samples of TB patients against tn2FbpC1-tnPstS1 showed 72.2% sensitivity which is nearly the same as the expected combined value for the two individual antigens. However, the sensitivity of tn1FbpC1-tnPstS1 was lowered to 60%. tnHSP-tn1FbpC1 showed 67.7% sensitivity which is slightly less than the expected combined value for the two individual antigens, but still significantly higher than that of each of the individual antigen. Data for secondary structure analysis by CD spectrometry was in reasonable agreement with the X-ray crystallographic data of the native proteins and the predicted structure of the fusion proteins. Comparative molecular modeling suggests that the epitopes of the constituent proteins are better exposed in tn2FbpC1-tnPstS1 as compared to those in tn1FbpC1-tnPstS1. Therefore, removal of the N-terminal non-epitopic region of FbpC1 from 34-96 amino acids seems to have unmasked at least some of the epitopes, resulting in greater sensitivity. The high level of sensitivity of tn2FbpC1-tnPstS1 and tnHSP-tn1FbpC1, not reported before, shows that these fusion proteins have great potential for use in serodiagnosis of tuberculosis.

KW - Epitopes

KW - FbpC1

KW - Fusion antigens

KW - HSP

KW - PstS1

KW - Serodiagnosis

UR - http://www.scopus.com/inward/record.url?scp=84930946131&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84930946131&partnerID=8YFLogxK

U2 - 10.1016/j.mimet.2015.06.003

DO - 10.1016/j.mimet.2015.06.003

M3 - Article

VL - 115

SP - 104

EP - 111

JO - Journal of Microbiological Methods

JF - Journal of Microbiological Methods

SN - 0167-7012

ER -