Complete nucleotide sequence of the Plasmodium vivax 6 kb element.
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Amino Acid Sequence↗
Biomedical subjects
Y D Sharma
Publications and source records attributed to Y D Sharma.
Alu elements in a Plasmodium vivax antigen gene.
Plasmodium vivax is a very common human malaria parasite but it is poorly characterized at the molecular level. Here, we describe the isolation and characterization of an antigen coding gene of P. vivax which contains Alu elements. This gene, called Pv-Alu, is expressed during the erythrocytic phase of the parasite. The encoded 200 amino acid long polypeptide is highly hydrophobic, contains transmembrane domains, and is rich in leucine (19.4%), serine (15.9%), proline (15.4%) and phenylalanine (15.4%). The 5'-untranslated region and part of the 3'-end coding region of Pv-Alu show significant homology to different Alu families. The presence of Alu elements in the coding region of a parasite antigen gene is significant from a functional and evolutionary viewpoint.
Amino Acid Sequence↗
Partial sequence analysis of a Plasmodium vivax cation transporting ATPase gene homologue & a putative pseudohomologue.
Molecular characterization of P. vivax is essential to develop suitable antimalarial drugs and vaccines. We describe here isolation and sequence analysis of a partial cDNA of a calcium ATPase as well as a putative pseudogene from this parasite. The immunoscreening of lambda gtll- P. vivax DNA library with patients serum has earlier resulted in the isolation of several seroreactive clones including Pv14. This clone contains a 299 bp insert having 18 amino acids (aa) reading frame fused with beta galactosidase. A larger fragment of approximately 15 kb was isolated from the EMBL3 library for Pv14 but it had only 2 extra aa in its reading frame. The far upstream region of Pv14 revealed a 101aa long putative open reading frame (ORF) showing homology to a variety of calcium ATPases in the M8 and M9 transmembrane region. But in the absence of a transcript in the parasite could indicate that it represents a pseudogene. However, the real gene for calcium ATPase in P. vivax was detected by RT-PCR using degenerate primers, designed from the conserved sequences of energy transduction and phosphorylation domains. The amplified cDNA-PCR product of 550 bp was cloned and sequenced which showed a significant aa homology to the calcium ATPase4 of P. falciparum. The present study, therefore, establishes the existence of calcium ion pumps in P. vivax which will be useful in drug development.
Amino Acid Sequence↗
Identification of a Plasmodium vivax heat-shock protein which contains a metalloprotease sequence motif.
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Amino Acid Sequence↗
Variations in the C-terminal repeats of the knob-associated histidine-rich protein of Plasmodium falciparum.
The knob-associated histidine rich protein (KAHRP) of Plasmodium falciparum plays an important role in the pathophysiology of cerebral malaria. In the present study, the immunogenic C-terminal repeat domain of the KAHRP gene was amplified, cloned and sequenced from the Indian (RJ181) and Honduran (HB3) isolates of P. falciparum. Based on the number and types of repeats in the domain, we report here the presence of three unique variant forms of KAHRP among these isolates. The Indian isolate (RJ181) contained four units of the decapeptide repeats whereas the Honduran isolate (HB3) contained two forms i.e. one form containing four decapeptide repeats plus a tetrapeptide subunit and the other form containing three decapeptide repeats plus a tetrapeptide subunit. Thus, all together, the number of KAHRP variants is increased to five which includes previously described two variants, each containing either 3 or 5 decapeptide repeats. This high rate of variability in the antigenic domain of the KAHRP gene via deletion or addition of whole or part of the decapeptide units could be involved in the evasion of host immune system possibly by providing the speculative complementarity to the vargene product. The results of the present study will be useful in designing the suitable molecular therapeutic reagents for cerebral malaria.
Amino Acid Sequence↗
Knob proteins in falciparum malaria.
Knob proteins play a significant role in the pathophysiology of cerebral malaria caused by Plasmodium falciparum. Most of these proteins are of parasite origin and can be divided into two major classes: (i) the cytoadherent proteins present at the surface of the knobs; and (ii) the submembranous structural proteins which are placed towards the cytoplasmic side in the knobs. Several surface proteins [viz., P. falciparum-infected erythrocyte membrane protein-1 (PFEMP-1), sequestrin, pfalhesin] and submembranous structural proteins [viz., knob-associated histidine-rich protein (KAHRP), PFEMP-2, PFEMP-3] of the knobs have been identified and characterized to a certain extent. The structural proteins interact with several host (e.g., spectrin, actin, band 4.1 etc.) as well as parasite (e.g., PFEMP-1) molecules to produce functional knobs. The surface proteins on the other hand interact with several adhesion molecules of the endothelial cell through receptor-ligand type of binding. Knob proteins are important from the point of view of malaria control since immunotherapeutic agents can be developed to block as well as reverse the cytoadherence phenomenon. The surface proteins are also good vaccine candidates except that they show a high rate of antigenic variation. Nevertheless, the use of ribozyme or antisense oligonucleotides to inhibit the expression of knob proteins (e.g., KAHRP alone or with surface protein) can be used as a molecular therapeutic agent.
Animals↗
High prevalence of chloroquine resistant Plasmodium falciparum infection in Rajasthan epidemic.
Plasmodium falciparum is the main killer among all human malaria parasites. In 1994, there was a falciparum malaria epidemic in Rajasthan, India, with many deaths. We have investigated active falciparum malaria cases from this epidemic and found that most of the parasite isolates (95%) were resistant to chloroquine. Nevertheless, all the tested isolates from the epidemic, were sensitive to mefloquine and quinine and ninety percent were also susceptible to sulfadoxine/pyrimethamine. Most individuals had moderate levels of TNF-alpha (20-220 pg/ml) and anti-parasite IgM antibodies compared to IgG levels which were relatively lower. In conclusion, the high transmission rate of the chloroquine resistant P. falciparum parasite could be the probable cause of the disease epidemic in Rajasthan. The timely drug sensitivity test and availability of appropriate antimalarial drugs are, therefore, warranted.
Animals↗
Allelic forms of the knob associated histidine-rich protein gene of Plasmodium falciparum.
The knob associated histidine-rich protein (KAHRP) gene was cloned and sequenced from two Indian isolates of Plasmodium falciparum, Pf3-92 and Pf29-92. These isolates showed major sequence differences in the C-terminal repeat domain of KAHRP. However, the biologically important domains such as spectrin-actin binding region remained highly conserved. The PCR amplification of a variable C-terminal repeat domain from the clinical isolates of P. falciparum, from Rajasthan epidemic, showed the presence of multiple alleles of KAHRP gene. The presence of multiple alleles indicates the existence of several P. falciparum strains in India. This should be taken into account for future malaria control strategies such as molecular therapy and vaccines.
Alleles↗
Enhanced expression of Plasmodium falciparum heat shock protein PFHSP70-I at higher temperatures and parasite survival.
The effect of various body temperatures, encountered during malaria fever, on the synthesis of Plasmodium falciparum heat-shock protein called PFHSP70-I and parasite growth rates among five different isolates are described. The results show that after the exposure of parasites at 39 degrees C for 30 min the amount of PFHSP70-I in all five isolates increased markedly and significantly, whereas parasite growth rates and the amount of total blood stage antigens remained almost unaffected. This indicates that the PFHSP70-I gene responds to heat-shock by producing higher amounts of PFHSP70-I protein, presumably to protect the parasite from being killed during malaria fever.
Animals↗
Plasmodium vivax: immune responses in a cross-section of the population in the Delhi area of India.
The immune status of the individuals living in the Delhi area of northern India where Plasmodium vivax is predominant is described. The majority of the individuals who were acutely infected with P. vivax had antibodies against crude blood-stage antigen and the recombinant antigen, PV9. In contrast, the peripheral blood mononuclear cells of the majority of these individuals did not show proliferation in the presence of either of these antigens. There was no correlation between antibody levels and T cell proliferation. Such an immune status of individuals living in endemic areas should be taken into account in the future in developing P. vivax malaria vaccines.
Adolescent↗
Effect of heat-shock on Plasmodium falciparum viability, growth and expression of the heat-shock protein 'PFHSP70-I' gene.
Cultures of the human malaria parasite Plasmodium falciparum were subjected to heat-shock for varying times and temperatures and then tested for their viability, growth and expression of heat-shock protein. Results show that the majority of parasites remained viable after heat-shock but their growth was affected. However, the expression of the heat-shock protein 'PFHSP70-I' gene was enhanced after heat-shock. We conclude that malarial parasites are able to survive in vivo during fever probably due to the overexpression of the heat-shock protein gene.
Animals↗
Cloning and developmental expression of the alpha 3 chain of chicken type IX collagen.
Fibrous and nonfibrous collagens comprise two major groups within the collagen family and both groups are found in a diverse variety of tissue fabrics. Type IX collagen is in the nonfibrous group; three different subunits of type IX collagen have been identified and the alpha 1 and alpha 2 subunits have been cloned. Using molecular cloning methods we have isolated, from an embryonic chicken cartilage library, cDNA clones which code for the entire alpha 3 chain of chicken type IX collagen. The cDNA clones encompass 2416 base pairs which have a conceptual open reading frame for a protein containing 675 amino acids including 193 Gly-X-Y repeats. These collagen repeats are in three separate domains which are interspersed with four major noncollagen domains. The collagen repeats also have four minor interruptions. This chain organization directly aligns with both the alpha 1 and alpha 2 chains of chicken type IX collagen. Comparison of the deduced amino acid sequence with peptide sequences of type IX collagens shows identity with 95 of the 96 known residues of the chicken alpha 3 chain and 81 of the 98 known residues of the bovine alpha 3 chain. The identical residues match those in five peptide fragments, two from the bovine protein and three from the chicken protein. The chicken and bovine alpha 3 chains have conserved cross-linking sites, separated by 137 residues which span 40 nm, the length of the hole zone in a collagen fibril. The NC3 domain of the chicken alpha 3 chain contains a repeat Cys-Pro motif which is present in both vertebrate and invertebrate nonfibrillar collagens. Northern blot hybridization exhibits a major mRNA of about 3.3 kilobases; this transcript is found in cartilaginous tissues in the embryo, including the developing limb and is not detected in other tissues or in the precondensation stage of limb development. The composite data delineate the primary structure of the alpha 3 chain of chicken type IX collagen, show its close relationship to the alpha 1 and alpha 2 chains, demonstrate its mRNA transcript, and show the appearance of that transcript in tissues of the developing chick embryo.
Amino Acid Sequence↗
Molecular cloning and serological characterization of a new Plasmodium vivax recombinant antigen which contains apolipoprotein B-100 like sequences.
We have isolated a new recombinant antigen 'PV12' of Plasmodium vivax by immunological screening of the genomic library. The antigen shows a sequence homology with human apolipoprotein B-100. A large number of P. vivax infected individuals (79%) were seropositive against this antigen. The probable function of this antigen could be to evade the host defence system (i) by inactivating the perforin of natural killer cells and (ii) by engaging the host immune system to produce antibodies against this decoy antigen.
Amino Acid Sequence↗
Structure and possible function of heat-shock proteins in Falciparum malaria.
Like many prokaryotes and eukaryotes, the malaria parasite also synthesizes several stress proteins. Most widely studied stress proteins of this parasite are the heat-shock proteins (hsps). Their discovery in malaria is a gift of recombinant DNA technology. Five hsp genes from Plasmodium falciparum have been identified which are located on different chromosomes. Thus the inheritance and expression of hsp genes are independent of each other. They share a large amount of sequence homology at N-terminus with the hsps of other organisms. Their gene regulatory sequences and other elements, important for gene expression, are yet to be determined. The biological role of these proteins in malaria is not fully understood but it is possible that they provide protection to the parasite from various stresses encountered in the host. In this process hsps probably bind to the toxic molecules as well as damaged proteins to flush them out of the parasite. Their involvement in the stage-specific parasite transformation to increase the infectivity and virulence, as observed in other parasites, remains to be determined. Malarial hsps are antigenic in humans. This antigenicity could be attributed to the non-homologous sequences in the C-terminus region. The potential of one of them (pfhsp 70I) for a future malaria vaccine and immunodiagnostics requires re-evaluation of the data.
Amino Acid Sequence↗
Recombinant fusion protein identified by lepromatous sera mimics native Mycobacterium leprae in T-cell responses across the leprosy spectrum.
Pooled polyvalent sera from lepromatous leprosy patients were used to screen a lambda gt11 recombinant DNA expression library of Mycobacterium leprae in order to identify the relevant antigens recognized by the human immune response. Of the 300,000 phages screened, 4 clones were identified that coded for fusion proteins of the same molecular mass. The fusion protein from clone LSR2 was tested for immunoreactivity in assays using peripheral blood cells and sera from 11 laboratory personnel and 105 patients across the leprosy spectrum. LSR2 protein appears to be predominantly a T-cell antigen. It evokes similar lymphoproliferative responses as the native bacillus both at the individual level and in the leprosy spectrum as a whole. Though only 50% of patient sera with anti-M. leprae antibodies reacted with the fusion protein, the pattern of reactivity in the antibody responses was also similar for the various clinical types. The coding regions of clones LSR1 and LSR2 are identical. They show no homology with sequences stored in data banks and encode a protein of 89 amino acids with a calculated molecular mass of approximately 10 kDa.
Amino Acid Sequence↗
Knobs, knob proteins and cytoadherence in falciparum malaria.
1. The sequestration of trophozoite and schizont infected erythrocytes (IRBC) in post-capillary venules of host internal organs causes most of the morbidity and mortality in falciparum malaria. It is a knob mediated cytoadherence phenomenon where knobs act as the focal junction between IRBC and host endothelial cell. Knobless (K-) parasites, isolated from cultures (not yet isolated from in vivo), do not cause virulent infections. Knobs thus play an important role in pathophysiology of falciparum malaria. 2. The chemical composition of knobs is partly explored, several proteins (Known as knob proteins) have been identified. According to their function they can be classified as (a) knob-inducing protein, "KAHRP" (b) knob-associated cytoadherent proteins, e.g. PFEMP-1, modified band 3 and an antigen recognized by monoclonal 33G2 and (c) knob-associated structural protein, e.g. PFEMP-2/MESA/PP-300. Most of them show size polymorphism among different isolates. Only KAHRP and MESA/PFEMP-2 have been studied at molecular level. Their chromosomal locations have been identified such as KAHRP on chromosome 2 and MESA/PFEMP-2 on chromosomes 5 and 6. 3. The receptor molecules on endothelial cells for knob ligands have been identified and partially characterized. 4. Knob ligands and their receptor molecules can play an important role in developing the immunotherapeutic reagents. 5. Based on the available data a tentative hypothesis has been proposed about the loss of knobs in vitro. Nevertheless, this needs further support from other experimental evidence. 6. Future work should be directed towards the structure and function of knob proteins and their interactions with each other as well as with host proteins. Regulation of expression of knobs and knob protein(s), evaluation of knob antigens for immunotherapy of severe falciparum malaria and for a malaria vaccine also require further investigations.
Amino Acid Sequence↗