Application and Development of Structural Vaccinology in Vaccine

Molecular Design Steps

Haoyan Jiang

College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei, China

Keywords: Structural Vaccinology, Vaccine Design, Antigenic Epitopes.

Abstract: Structural vaccinology has a broad prospect, which takes genomic research as a basis, combines research

directions from multiple disciplines, and advocates the use of surface proteins or three-dimensional structures

of pathogens to guide vaccine design. However, there is still a lack of systematic knowledge about structural

vaccinology's specific mechanism and application value in vaccine molecular design. This paper provides a

brief introduction to structural vaccinology and the molecular design of vaccines, a review of the application

of structural vaccinology in molecular design, an in-depth discussion of the role of structural vaccinology in

the determination of antigenic epitopes, epitope synthesis and vector construction, and finally a brief overview

of the current status and prospects of structural vaccinology. By collecting and analysing relevant data, this

paper discusses in depth the application strategies, technological progress and future development direction

of structural vaccinology in the key aspects of vaccine molecular design to provide new ideas and

methodological references for vaccine research and development.

1 INTRODUCTION

A vaccine is a biological product that induces the host

to develop immune resources against a particular

antigen, interrupting the transmission of an infectious

agent while helping the host to prevent infection.

Vaccines also enhance the specific immunity of the

inoculated population against an antigen, which is

effective in preventing disease and treating it. The

invention of vaccines is considered one of the

triumphs of medical research. Immunisation stops the

spread of infections in childhood and provides

lifelong protection against certain diseases.

Currently, the four types of vaccines available to

humans are inactivated, live attenuated, subunit and

nucleic acid vaccines. Live attenuated vaccines

include the whole pathogen, which can replicate in

the host and induce a strong immune response.

Whole-pathogen inactivated vaccines are mostly safe

and non-infectious and generally inactivated by

physical heating and many chemical methods.

Inactivated vaccines have now been tested and found

to be deficient in inducing weak and short-term

immunity and, therefore, require boosters to enhance

their immune effect to achieve complete protection.

Subunit vaccines contain purified or recombinant

antigens consisting of only the antigenic portion of

the pathogen, not the entire cell. They are, therefore,

generally safe regarding toxicity and reactogenicity

(Verma et al. 2023).

Structural vaccinology, as a discipline focusing on

exploring the structure-function relationship of

vaccines, plays a key role in understanding the

working principle of vaccines and promoting the

optimisation of vaccine design. With the help of X-

ray crystal diffraction, cryo-electron microscopy and

other cutting-edge structural biology techniques,

researchers can precisely analyse the structure of

vaccines and gain in-depth insights into the intrinsic

mechanisms of vaccine-induced immune responses,

thus building a solid theoretical foundation for the

design and development of new vaccines. In addition,

through computer-aided simulation and prediction,

the interactions between vaccines and immune cells

can also be studied, thus providing a more thorough

understanding of the working mechanism of vaccines

and helping to realise breakthroughs and

developments in vaccinology.

After a long period of scientific research,

analysing the three-dimensional structures of most

protein antigens at the atomic level is now possible,

thanks to the development of structural biology

techniques. In addition, in-depth studies of structural

198

Jiang, H.

Application and Development of Structural Vaccinology in Vaccine Molecular Design Steps.

DOI: 10.5220/0014446300004933

Paper published under CC license (CC BY-NC-ND 4.0)

In Proceedings of the 1st International Conference on Biomedical Engineering and Food Science (BEFS 2025), pages 198-202

ISBN: 978-989-758-789-4

biology have provided important structural

information that can guide design modifications at the

atomic level. In particular, the structures of antigen-

protective monoclonal antibody complexes have

revealed key epitopes for antigen recognition, which

has led to a deeper understanding of the mechanisms

of the host's protective immune response and thus

guided the reverse design of vaccines. The structural

biology-based vaccine design approach is a new

research direction (Li et al. 2024). The development

of synthetic vaccines can avoid the use of attenuated

and inactivated disease-causing pathogens, further

reducing the risks associated with their pathogenicity.

These vaccines can be designed to trigger specific

immune responses more precisely.

As an emerging field of vaccine research and

development, the development of structural

vaccinology has provided an important theoretical

basis and technical support for enhancing vaccine

effectiveness and promoting innovative

breakthroughs in vaccine design technology. In this

paper, we systematically integrate the research

progress of structural biology in vaccine molecular

design in recent years and discuss the basic principles,

key technical paths and specific implementation steps

of vaccine molecular design, aiming to enrich the

research connotation of vaccine molecular design

from the theoretical level and to provide scientific

references and bases for the optimisation of vaccine

molecular infrastructures and the improvement of

vaccine efficacy.

2 DEFINITION AND

MECHANISM OF ACTION OF

VACCINES

Vaccines are automatic immunising agents for

medical use. They are generally divided into two

categories: prophylactic vaccines, which generally

act on healthy individuals such as newborns, and

therapeutic vaccines, which generally act on diseased

individuals. According to tradition and custom, there

are also types at the molecular level, such as live

attenuated vaccines, inactivated vaccines, antitoxins,

subunit vaccines (including peptide vaccines), vector

vaccines, and nucleic acid vaccines.

The principle of action of vaccines is to treat

pathogenic microorganisms and their metabolites

using methods such as genetic engineering or

artificial manipulation to reduce virulence so that they

retain their properties of stimulating the immune

system of the animal body. The immune system will

produce certain protective substances, such as

immune hormones, active physiological substances,

unique antibodies, etc., when the animal body is

exposed to such attenuated or inactivated pathogenic

bacteria. When the protective cells in the animal's

immune system recognise the antigen again, they will

be stimulated to produce an immune response and

make the same antibodies to prevent the pathogenic

bacteria from harming the animal, which ultimately

leads to immune protection of the body.

3 MOLECULAR DESIGN OF

VACCINES

The molecular design of vaccines refers to the in-

depth analysis of the structure of vaccines and their

immune effects through modern molecular biology

and immunology techniques, then design and

optimise them to make vaccines safer and more

efficient. The design concept is to identify the key

components in the molecular structure of the

pathogen that triggers the immune response and

modify them according to the characteristics of its

biology (Li et al. 2024). The development of synthetic

vaccines can avoid attenuated disease-causing

pathogens and reduce the risks they pose. At the same

time, these vaccines can be designed to trigger

specific immune responses more precisely.

The molecular design of vaccines encompasses a

variety of techniques. The first is using computer-

based simulation programs to predict pathogen

proteins' spatial conformation and three-dimensional

structure, providing strong evidence for the design of

vaccine molecules that match them. The second is

screening effective vaccine molecules through

multiple comparative testing experiments. The third

is to utilise bioinformatics tools and algorithms to

design efficient vaccine molecules. The fourth is to

design chimeric vaccine molecules by combining the

principle of high structural similarity of different

types of surface antigens. Fifthly, chemical synthesis

and bioengineering techniques are used to assemble

and modify antigens and other molecules so that they

have vaccine functions. In conclusion, the molecular

technology of vaccines requires an in-depth study of

pathogens' structure and biological characteristics. It

is a highly critical and complex process in vaccine

development.

Application and Development of Structural Vaccinology in Vaccine Molecular Design Steps

199

4 STRUCTURAL

VACCINOLOGY AND ITS

APPLICATION TO VACCINE

MOLECULAR DESIGN

4.1 The Concept of Structural

Vaccinology and Its Development

The concept of structural vaccinology (SV) was

introduced in 2008, advocating the study of utilising

surface proteins or three-dimensional structures of

pathogens to guide vaccine design, and this research

direction is considered to have a broad prospect.SV

takes genome study as a foundation and combines the

research directions of multidisciplinary fields such as

molecular biology and structural biology. By utilising

techniques such as X-ray crystallography, nuclear

magnetic resonance spectroscopy and cryo-electron

microscopy, researchers can dissect the molecular

structure of pathogen antigens and further understand

the mechanisms by which they interact with the host

immune system (Li et al., 2024).

SV research suggests that machine learning

algorithms rarely augment SV and are used as an aid.

A potential extension of SV design is the use of

evolutionary algorithms that evaluate assessments

and replacements based on multiple rounds of data

evaluation, provide variables by modifying inputs,

and optimise and adjust for the function's results to

improve its fitness after several iterations have been

performed. However, there is still a lack of good

scoring functions to ensure that the evolutionary

algorithm will produce optimised vaccine candidates

rather than maximising some arbitrary mechanism

(Huffman et al. 2022).

4.2 Application of Structural

Vaccinology to Specific Steps of

Vaccine Molecular Design

4.2.1 Determination of Antigenic Epitopes

Antigenic epitope determination identifies key

antigenic epitopes in viral proteins by molecular

biology methods. African swine fever (ASF) is highly

infectious and lethal, and its causative agent is the

African swine fever virus (ASFV). Only in-depth

studies of ASFV antigens can facilitate the

development of effective vaccines and control

measures. p22 protein, one of the major structural

proteins of ASFV, can be detected at the early stages

of ASFV infection, making it a potential candidate

protein for detecting ASFV(A et al. 2021).To carry

out the preliminary identification of antigenic

epitopes recognised by monoclonal antibodies,

firstly, it is necessary to use PSIPRED software to

predict the secondary structure of the p22 protein and

get that the protein is composed of four α-helices and

seven extended chains; secondly, eight peptides were

designed and synthesised according to the amino acid

sequence and secondary structure of p22 protein, and

Dotblotting preliminarily identified the antigenic

epitopes of p22 protein. Then, TMHMM-2.0 software

was used to predict the transmembrane structural

domains of the protein and the tertiary structure of the

p22 protein; finally, antigenic epitope analysis was

carried out, and the antigenic epitopes obtained by

identification were compared with the amino acid

sequences of the p22 protein of 20 different ASFVs

in GenBank. The results showed that the sequences of

the two antigenic epitopes recognised by the

monoclonal antibody were highly conserved, and

there was no difference in amino acid sequences

(Wang et al. 2025).

4.2.2 Epitope Synthesis

Epitope synthesis uses synthetic peptide technology

to synthesise these antigenic epitopes artificially, and

its methods include two major categories: chemical

synthesis and biosynthesis. Chemical synthesis is

divided into solid-phase synthesis and liquid-phase

synthesis. Solid-phase synthesis is the step-by-step

construction of peptide chains by connecting amino

acids to solid-phase carriers one by one, usually used

for the synthesis of short peptides, which is the most

commonly used method of peptide synthesis; liquid-

phase synthesis is carried out in solution, usually used

for the synthesis of longer peptide chains with

complex spatial structures. The biosynthesis method

also consists of two methods: one is edited using

genetic engineering technology, inserting gene

fragments encoding antigenic epitopes into

expression vectors, and finally extracting the target

products in host cells so that epitopes with natural

conformations can be synthesised; the other is to

clone the gene fragments encoding polypeptides and

introduce them into phages so that the final translated

polypeptides are presented on the surface of the

phages In the other case, the gene fragment encoding

the peptide is cloned and introduced into the phage so

that the final translated peptide is presented on the

surface of the phage.

Helicobacter pylori (Hp) is a Gram-negative

bacterium typically found in the gastric epithelium of

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humans (Xu et al. 2021).In 2022, a study was

conducted to synthesise the core undecanoate, outer

core pentasaccharide, outer core pentasaccharide,

inner core trisaccharide, phosphorylated inner core

trisaccharide, and α-1,6-glucan in the structure of the

Hp lipopolysaccharides using a chemical method, and

the antibody affinity of the synthesised

oligosaccharides was evaluated using a glycan chip

technique. The experimental results showed that α-

1,6-glucan could bind well to serum IgG antibodies

of most Hp-infected patients, and this study

demonstrated that α-1,6-glucan may be an important

antigenic epitope of Hp lipopolysaccharide (Zou et al.

2022). A series of oligosaccharide molecules formed

from monosaccharide units linked by glycosidic

bonding were first generated by a catalytic reaction of

tens of steps using tribenzyl oxidised boron benzoate

(Zhao et al. 2020) as a starting material from

monosaccharides to pentasaccharides, with the yields

of each molecule being 90%, 88%, 90%, 92%, and

89%, respectively. Each sugar unit is a pyranose ring

structure and carries a hydroxyl group, and the

pentasaccharide molecule carries an amino group at

the end (Zhao et al. 2024).

4.2.3 Carrier Construction

To enhance immunogenicity, vector construction is

the insertion of synthetic antigenic epitopes into

appropriate vectors, such as DNA vaccines or virus-

like particles. In order to prevent and reduce losses, a

study utilised Red homologous recombination

technology, using bacterial artificial chromosome

(BAC) as a gene editing platform, to integrate the F

gene of NDV into the genome of MDV double

deletion strain Md5BACΔmeqΔLorf9, thereby

knocking out the F gene of NDV and inducing I-SceI

enzyme expression. The F gene of NDV was

integrated into the genome of MDV double deletion

strain Md5BACΔmeqΔLorf9, which induced the

expression of the I-SceI enzyme, thereby knocking

out the kanamycin resistance gene, resulting in the

successful construction of the recombinant live-

vector vaccine candidate strain

Md5BACΔmeqΔLorf9-F (Gong et al. 2024). In order

to construct a recombinant adenovirus with the

replication-defective human adenovirus type 5 (Ad5),

the capsid protein of duck tambuusu virus (DTMUV),

another study inserted the DTMUV Capsid gene into

the pShuttle-CMV-Hsp70 plasmid containing the

heat shock protein 70 (mHsp70) of Mycobacterium

tuberculosis as the adjuvant by a one-step cloning

technique and constructed a recombinant adenovirus

able to The DTMUV Capsid gene was inserted into

the pShuttle-CMV-Hsp70 plasmid containing

Mycobacterium tuberculosis heat shock protein 70

(mHsp70) as an adjuvant, to construct the

recombinant shuttle plasmid pShuttle-DTMUV

Capsid, which can express DTMUV Capsid and

mHsp70 proteins (Wu et al. 2024).

5 CONCLUSION AND OUTLOOK

This paper summarises the definition of a vaccine and

its principle of action, as well as describes the

concept, application, and prospects of structural

vaccinology. Structural vaccinology plays a pivotal

role in many steps of vaccine molecular design, which

can not only determine antigenic epitopes by

predicting and comparing the secondary and tertiary

structures of proteins but also artificially synthesise

antigenic epitopes by using synthetic peptide

technology, which helps to construct carriers for

antigenic epitopes to enhance immunogenicity.

However, although structural vaccinology has

made certain breakthroughs and development, it still

faces serious challenges: first, the difficulty of

structural analysis, some antigens are difficult to

crystallise or complex structure, which limits its

application; second, the pathogen will escape the

immune response through mutation, resulting in

immune escape phenomenon, which makes the

vaccine effect decline; third, the high cost of

structural vaccinology technology, which restricts its

large-scale applications Thirdly, the high cost of

structural vaccinology technology limits its large-

scale application, and how to reduce the cost is also

an urgent problem to be solved.

Vaccine design methods based on structural

biology have become a promising research direction.

In vaccine production and supply, innovations in

vaccine molecular design will not only reduce costs

and increase efficiency but also promote the

improvement of vaccine production technology,

which on the one hand, will enable the development

of broad-spectrum vaccines capable of responding to

a wide range of pathogen variants, and on the other

hand will enable the design of personalised vaccines

based on the differences in the individual's immune

system, so that the efficiency of the vaccine supply

can be increased rapidly to meet the needs of global

public health. In response to new outbreaks, the

increasingly mature vaccine molecular design

technology can rapidly analyse the pathogens and

design targeted vaccines at the early stage of an

Application and Development of Structural Vaccinology in Vaccine Molecular Design Steps

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outbreak, and combined with the development of new

adjuvants and delivery systems, it can improve the

immunogenicity and stability of vaccines and achieve

rapid prevention and control of infectious diseases. In

addition, with the continuous development of

structural biology technology, the precision and

efficiency of 3D structure analysis of antigenic

proteins will continue to improve. Based on the high-

resolution structural information, the

immunogenicity and specificity of antigens can be

improved through strategies such as antigen epitope

modification and protein multimerisation design,

laying a solid foundation for developing new-

generation vaccines.

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