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Antibody Binding Epitopes of an NTHi Moonlighting Protein

Alvarado, Vera (2016) MOBT01 20152
Degree Projects in Molecular Biology
Abstract
After the introduction of a conjugate vaccine that provides immunization to Haemophilus influenzae type b, an increase in the number of infections caused by Non-typeable H. influenzae (NTHi) has been reported. NTHi mostly affects children under the age of 1, causing cases of otitis media and conjunctivitis, and it is also associated with respiratory tract infections such as exacerbations of chronic obstructive pulmonary disease affecting adults over 65 years. NTHi possesses several strategies for infection and evasion of the host immune system, surface proteins play an important role in this line of attack. In recent proteomic studies, the protein Elongation Factor Thermo unstable (EF-Tu) has been shown to be present in the outer membrane... (More)
After the introduction of a conjugate vaccine that provides immunization to Haemophilus influenzae type b, an increase in the number of infections caused by Non-typeable H. influenzae (NTHi) has been reported. NTHi mostly affects children under the age of 1, causing cases of otitis media and conjunctivitis, and it is also associated with respiratory tract infections such as exacerbations of chronic obstructive pulmonary disease affecting adults over 65 years. NTHi possesses several strategies for infection and evasion of the host immune system, surface proteins play an important role in this line of attack. In recent proteomic studies, the protein Elongation Factor Thermo unstable (EF-Tu) has been shown to be present in the outer membrane of several bacterial species. Given this fact, characterization of human antibodies reactive to the protein might provide further understanding of its role as an emerging antigen determinant. This study was done to determine relevant immunogenic sites of NTHi EF-Tu. By using truncated fragments of recombinant EF-Tu, achieved by two different approaches (CNBr cleavage and expressing protein constructs containing different domains of the protein), human sera tested in this study showed binding to recombinant NTHi EF-Tu Domain 1, and CNBr-cleaved fragments encompassing a region from the C-terminus of Domain 1 to the N-terminus of Domain 3 of NTHi EF-Tu. (Less)
Popular Abstract
Acquired protection against Haemophilus influenzae

There are numerous ways in which microorganisms are able to evade the integrity of human natural barriers against invaders. However our body manages to produce proteins called antibodies that will recognize an attacker and trigger other proteins to eliminate it. How the body manages to produce such antibodies is a fascinating but exceedingly complex process that has been the subject of immunology, and studying this has helped to produce effective vaccines.

Haemophilus influenzae type b, was one of the leading causes of meningitis in children before the introduction of a vaccine. However the vaccine does not provide protection against other types of Haemophilus. In fact, although it... (More)
Acquired protection against Haemophilus influenzae

There are numerous ways in which microorganisms are able to evade the integrity of human natural barriers against invaders. However our body manages to produce proteins called antibodies that will recognize an attacker and trigger other proteins to eliminate it. How the body manages to produce such antibodies is a fascinating but exceedingly complex process that has been the subject of immunology, and studying this has helped to produce effective vaccines.

Haemophilus influenzae type b, was one of the leading causes of meningitis in children before the introduction of a vaccine. However the vaccine does not provide protection against other types of Haemophilus. In fact, although it is believed to be unrecognized, nowadays non-typeable H. influenzae (NTHi) is an important cause of airways infections in adults over 65 years and it also affects children under 1 year, causing otitis.

When a particular protein, necessary for the life of the bacteria or produced to make invasion easier, is present on the surface of the bacteria, an antibody can reach the bacteria easily, bind to it and destroy it. Originally thought to be inside the cell, the protein Elongation Factor Thermo unstable (EF-Tu) has been found to be present on the surface of several bacteria. As a result, antibodies against this protein are expected to be found. Furthermore, being able to recognize the identity of the binding site that leads to the production of antibodies gives additional insight for the development of possible candidates for vaccines. To the date, no information regarding the binding sites of human antibodies against EF-Tu has been described.

A particular region of EF-Tu reacts with human antibodies
By analyzing different parts of EF-Tu and sera from human donors, antibodies specific against the protein were found. EF-Tu has three different regions, called domains. These regions allow the protein to fulfill its main biological function, which is the production of proteins. Due to the latter, it is important to mention that EF-Tu is found with only minor differences in all bacteria. It does not vary much since all parts of the protein are needed for its function, reinforcing the necessity of determine the specific site of binding for the antibodies.

Two regions, one probably involving only Domain 1, and other involving parts of Domains 1, the entire Domain 2 and parts of Domain 3, showed to attract the immune system and elicit antibodies. However, more studies should be carried on to determine the reaction of human antibodies against EF-Tu of NTHi and other bacteria.


Advisors: Prof. Kristian Riesbeck, Dr. Ben Duell & Dr. Yu-Ching Su
MasterĀ“s Degree Project for 60 credits in Molecular Biology, Microbiology 2016
Department of Biology, Lund University (Less)
Please use this url to cite or link to this publication:
author
Alvarado, Vera
supervisor
organization
course
MOBT01 20152
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
8891803
date added to LUP
2016-09-15 09:47:59
date last changed
2016-09-15 09:47:59
@misc{8891803,
  abstract     = {After the introduction of a conjugate vaccine that provides immunization to Haemophilus influenzae type b, an increase in the number of infections caused by Non-typeable H. influenzae (NTHi) has been reported. NTHi mostly affects children under the age of 1, causing cases of otitis media and conjunctivitis, and it is also associated with respiratory tract infections such as exacerbations of chronic obstructive pulmonary disease affecting adults over 65 years. NTHi possesses several strategies for infection and evasion of the host immune system, surface proteins play an important role in this line of attack. In recent proteomic studies, the protein Elongation Factor Thermo unstable (EF-Tu) has been shown to be present in the outer membrane of several bacterial species. Given this fact, characterization of human antibodies reactive to the protein might provide further understanding of its role as an emerging antigen determinant. This study was done to determine relevant immunogenic sites of NTHi EF-Tu. By using truncated fragments of recombinant EF-Tu, achieved by two different approaches (CNBr cleavage and expressing protein constructs containing different domains of the protein), human sera tested in this study showed binding to recombinant NTHi EF-Tu Domain 1, and CNBr-cleaved fragments encompassing a region from the C-terminus of Domain 1 to the N-terminus of Domain 3 of NTHi EF-Tu.},
  author       = {Alvarado, Vera},
  language     = {eng},
  note         = {Student Paper},
  title        = {Antibody Binding Epitopes of an NTHi Moonlighting Protein},
  year         = {2016},
}