Apical Membrane Antigen (AMA1) is a protein expressed in the membrane
of Plasmodium falciparum, a protozoan parasite which causes the most
deadly form of malaria in humans. Malaria is an infectious disease which is
responsible for up to 2% of global mortalities. It kills up to 2.7 million
people each year, most of whom are children in Sub-Saharan Africa. Malaria
parasites are transmitted to humans via female Anopheles
When an infected mosquito bites a human, the disease enters the
hepatic, or exoerythrocytic phase, and Plasmodium sporozoites in
the mosquito’s saliva enter the bloodstream and migrate to the liver. Once
in the liver, the sporozoites infect hepatocytes and then multiply and differentiate
into thousands of merozoites. Following rupture of their host cells, the
merozoites move to the blood where they infect red blood cells; at this
point the disease enters the erythrocytic stage. The parasites continue
to multiply in the red blood cells, periodically lysing the cell to invade
new red blood cells. As the cycle continuous the malaria symptoms in the
individual progressively worsen
For an overview of the pathogenesis of malaria click here
Historically, malaria has been combated using preventative measures
and expensive drug treatments, which are not affordable to the majority
of the world's malarial population. Furthermore, many strains of Plasmodium
species have developed resistance to these drugs. Recently however, scientists
have identified AMA1 as a leading candidate for inclusion in a malaria vaccine.
AMA 1 is highly conserved in all apicomplexa parasites and is
expressed in the merozoite stage of the parasite's life cycle. Experimental
evidence has shown that AMA1 is involved in merozoite invasion of red blood
cells, and that is essential to the proliferation and survival of the malarial
parasite. More importantly, by introducing specific anti-AMA1 antibodies into
infected cell cultures, studies have shown that malaria parasites are unable
to infect red blood cells, and thus unable to reproduce and spread disease.
This tutorial provides the structures of domain I and II of AMA1
(making up amino acid residues 108-438) as they were crystallized by Bai
(2005), and discusses the molecular bases for AMA1 red blood cell binding,
evolution of resistance to human antibodies, and vaccine development.
II. General Structure
The extracellular region of AMA1 consists of three domains, the
first two of which, domain I and domain
II make up the central and most conserved portion of AMA1
. These regions were crystallized by Bai (2005) and are shown interacting
in the figure to the left. A 20 amino acid, conserved, N-terminal
sequence domain extending from domain I, was also crystallized. This
extension forms a distorted helix that lies against domain II
In domain II, a loop containing an alpha-helix and a pair of beta-strands
extends outwards from its center
. Other than this loop, domain II forms a typical protein domain, consisting
mostly of secondary structures which interact hydrophobically. Domain I on
the other hand, is fairly irregular, consisting largely of non repetitive
structure, and six loop extensions
At the core of both domains lies a conserved PAN
consisting of a central five-stranded beta-sheet
, which wraps around a 12- residue alpha-helix
. The AMA1 PAN domains are relatively exceptional when compared to PAN domains
of other proteins (Bai, 2005). For instance, only one disulfide
and six hydrophobic residues
are conserved within each PAN domain. In fact, since domains I and II lack
certain characteristic disulfide bonds and have acquired others, they were
first not recognized as PAN domains. Furthermore, Loops connecting beta strands
1 and 3, and
4 and 5 adopt variable conformations, and
the beta strands themselves, are variable in length and position
It has been suggested that these irregularities with the PAN domains,
particularly the flexible loops connecting the beta-sheets, resulting from
AMA1 evolution, divert antibodies from functionally critical regions of the
PAN domain, providing the malaria parasite with a way to evade protective
III. Erythrocyte Binding
Eight Peptides have been identified for their high affinity binding to erythrocytes. Four are conserved throughout the Plasmodium species: peptides 4313 and 4321, located in domain I
, peptide 4325, located in domain II
, and 4337, located in the cytoplasmic domain which was not crystallized. All four of these peptides are important in maintaining the three dimensional peptide structure necessary for erythrocyte binding. Peptides 4313, 4321, and 4322
share similar sequences to the MAEBL erythrocyte binding domains of other
Plasmodium species. In addition, peptide 4322
has been very important for vaccine development because it shares a similar
amino acid sequence to a B-cell epitope, an antibody binding site, that
has significantly decreased growth of P. falciparum in candidate
vaccines (Urquiza, 2001).
Within P. falciparum AMA1 domain I, lie five highly polymorphic residues which form a cluster at positions 187, 197, 200, and 243 on domain I and 230 on domain II, on one side of AMA1
. Other high and low frequency dimorphic sites are also located on this side of AMA1, termed “the polymorphic face”.
Only four dimorphic residues lie on the nonpolymorphic side, two of which occur with low frequency
. To view an image comparing the polymorphic and nonpolymorphic faces of the molecule with both dimorphisms and polymorphisms illustrated click here (from Bai, 2005). Due to the stark contrast in clustering of polymorphisms on one side of AMA1, it has been supposed that this single face of the molecule is exposed on the parasite surface (Bai, 2005).
V. Invasion-inhibitory antibodies
The administration of particular antibodies to cells infected with malarian parasites has been shown to inhibit parasitic invasion of the host red blood cells, thus reducing spread of the disease. Here we describe the strain specific binding of a monoclonal antibody (MAb) 1F9 (Coley et. al., 2006), to a region of P. falciparum 3D7 AMA1 which overlaps with the binding site of other antibodies on particular strains of AMA1 (Harris et. al., 2005). MAb 1F9 binds to an AMA1 amino acid region 57 residues long (191-247) which lies within PAN domain I . Residue 197 (E in 3D7 AMA1), the most polymorphic residue of AMA1 with up to 7 polymorphisms, lies within this region, and any single mutation of this residue has resulted in abolished or significantly reduced binding affinity .
VI. Future Prospects
Invasion of red blood cells is essential to the life cycle of the malaria
parasite making it a potential target for designing a vaccine that would
prevent the multiplication of the parasite. The molecular interactions that
govern this process such as host cell binding, merozoite reorientation,
and junction formation, parasitophorous vacuole formation, and host cell
entry are only some of the areas scientists have just started to examine(
Howell et al. 2001). Scientists are studying a hydrophobic cleft in the
molecule that is suspected of being the site where red blood cells are tethered
to AMA1. Future research will concentrate on finding compounds that can
fit inside the trough and block the parasite from attaching to red blood
cells. As of June 2006, no completely effective vaccine is yet available.
Hopefully these molecular investigations will further the development of
a new AMA1 vaccine against malaria (Mgrdichian).
Bai, Tao, Becker, Michael, Gupta, Aditi, Strike, Phillip, Murphy, Vince J., Anders, Robin F., and Batchelor, Adrian H. 2005. Structure of AMA1 from Plasmodium falciparum reveals a clustering of polymorphisms that surround a conserved hydrophobic pocket. PNAS 102: 12736-12741.
Coley, A.M, K. Parisi, R. Masciantonio, J. Hoeck, J.L. Casey, V.J. Murphy, K.S. Harris, A.H> Batchelor, R.F Anders, and M. Foley. The most polymorphic residue on Plasmodium falciparum apical membrane antigen 1 determines binding of an invasion inhibitory antibody. Infection and Immunity 74: 2628-2636.
Chen, B., Vogan, E.M., Gong, H., Skehel, J.J., Wiley, D.D., and Harrison, S.C. 2005 Nature 433: 834-841.
Healer, Julie, Murphy, Vince, Hodder, Anthony N., Masciantonio, Rosella, Gemmill, Alan, W., Anders, Robin F., Cowman, Alan F., and Batchelor, Adrian. 2004. Allelic polymorphisms in apical membrane antigen-1 are responsible for evasion of antibody-mediated inhibition in Plasmodium falciparum. 52(1): 159-168.
Howell, Steven A., Chrislaine Withers- Martinex, Clemens H.M. Kocken,
Alan W. Thomas, and Michael J Blackman. 2001. Proteolytic processing and primary
structure of Plasmodium falciparum apical membrane antigen 1. The
Journal of Biological Chemistry 267: 31311-31320.
Mgrdician, Laura. 2005. A step closer to a malaria vaccine. Tropical Disease News. Brookhaven National Laboratory.
Skehel, J.J., and Wiley, D.C. 2000. Annual Revue of Biochemistry 69: 531-569.
Urquiza, Maurico, Suarez, Jorge E, Cardenas, Constanza, Lopez, Ramses, Puentes, Alvaro, Chavez, Francisco, Calvo, Julio Cesar, Patarroyo, Manuel Elkin. 2001. Plasmodium falciparum AMA-1 erythrocyte binding peptides implicate AMA-1 as erythrocyte binding protein. Vaccine 19: 508-513.
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