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B H Kalinna

Publications and source records attributed to B H Kalinna.

7 recordsLinked to original sources
PubMed1996

Cloning and characterization of a ribosomal P protein from Taenia solium, the aetiological agent of human cysticercosis.

A cDNA encoding the complete open reading frame of the Taenia solium acidic ribosomal phosphoprotein P2 has been cloned. The 417 bp cDNA (arpP2) was isolated from a T. solium cDNA expression library by PCR using P protein specific pimers. The ORF encodes a protein of 121 amino acids exhibiting 40-55% identity to mammalian, fungal, insect, and parasite P2 proteins. The inferred molecular mass of the protein is 12,592 Daltons, similar to that reported for other ribosomal P2 proteins. After subcloning and expression, the recombinant protein was purified by affinity chromatography under non-denaturing conditions and was shown to have a molecular mass of 15 kDa which is equivalent to the expected size of the full length recombinant fusion protein, comprising the 12.5 kDa arpP2 plus an additional 2 kDa for the N-terminal fusion peptide incorporating the six histidine residues required for purification.

Amino Acid Sequence↗
PubMed1995

Cloning and characterization of the Schistosoma japonicum aspartic proteinase involved in hemoglobin degradation.

A cDNA encoding a Schistosoma japonicum aspartic proteinase was cloned, sequenced, and found to encode a zymogen of 380 amino acid residues, and its gene was shown to be present as a single copy in the S. japonicum genome. Identity comparisons showed that the enzyme (Sjpasp) was most closely related to the cathepsin Ds. The deduced amino acid sequence has four potential glycosylation sites, two of which are in identical positions to the two glycosylation sites of human kidney lysosomal cathepsin D. Furthermore, all four disulfide bonds found in mammalian cathepsin D sequences are present in Sjpasp, although the beta-hairpin (loop 3), which is cleaved during maturation of vertebrate cathepsin Ds to yield light and heavy chain subunits, is absent from Sjpasp. While most residues involved in substrate specificity and catalysis of aspartic proteinases are preserved in Sjpasp, several residues in these regions exhibit changes that may result in a novel substrate specificity. Aspartic proteinase activity is present in extracts of adult S. japonicum and Schistosoma mansoni and in culture media in which schistosomes were maintained and was capable of digesting hemoglobin. The schistosome aspartic proteinase may play a pivotal role in the catabolism of hemoglobin obtained from host erythrocytes.

Amino Acid Sequence↗
PubMed1995

Schistosoma japonicum: heterogeneity in paramyosin genes.

Paramyosin is an integral muscle protein found in many invertebrates including schistosomes, and is considered an important candidate vaccine antigen in schistosomiasis. The characterisation of natural molecular variation in vaccine antigens including paramyosin is important because strain-specific vaccination may be necessary against schistosomiasis japonica. We have isolated partial cDNAs encoding paramyosin from an adult, Chinese strain Schistosoma japonicum gene library. Two of these cDNAs (B6 and Y6) encode the same region of paramyosin but their nucleotide sequences differ at eight positions and their deduced amino acid sequences differ by an arginine/cysteine substitution, demonstrating intrastrain variation in paramyosin. Southern blot analysis of genomic DNA from the Chinese and Philippine strains of S. japonicum demonstrated strain-related RFLPs at the paramyosin locus, and suggested that more than one copy of the paramyosin gene was present in the S. japonicum genome. PCR-based RFLP analysis which exploited restriction site differences between B6 and Y6 showed that paramyosin genotype B6 was much more common in schistosome populations and verified the existence of introns in the paramyosin gene(s) of S. japonicum.

Amino Acid Sequence↗
PubMed1995

Gene cloning and complete nucleotide sequence of philippine Schistosoma japonicum paramyosin.

The development of an effective vaccine is recognised as a necessary adjunct to the control of schistosomiasis japonica, a disease affecting several million people in China and the Philippines. Currently, recombinant Schistosoma japonicum molecules are considered most suitable for large scale vaccine production and a number of genes encoding vaccine candidate polypeptides have been cloned and expressed (see Waine et al., 1993a). One of the molecules providing most promise as a vaccine target is paramyosin (Butterworth, 1992), a major structural protein of thick filaments in the muscle of most invertebrates; paramyosin genes have now been cloned from a range of parasitic helminths, including schistosomes (Limberger and McReynolds, 1990; Laclette et al., 1991; Dahmen et al., 1993; Landa et al., 1993; Mühlschlegel et al., 1993, Nara et al., 1994). The cloning and nucleotide sequence of S. Japonicum paramyosin is described.

Animals↗
PubMed1994

Gene cloning, overproduction and purification of a functionally active cytoplasmic fatty acid-binding protein (Sj-FABPC) from the human blood fluke Schistosoma japonicum.

We report the gene cloning, molecular characterisation and purification of a 14.7-kDa functionally active recombinant (re) cytoplasmic fatty acid-binding protein (Sj-FABPC) from the Chinese strain of the human bloodfluke Schistosoma japonicum (Sj). As schistosomes are unable to synthesise long chain fatty acids and sterols de novo and must, therefore, take up these lipids from the host, Sj-FABPC is an attractive vaccine and/or drug target. Clone 39 (C39), which contains the entire Sj-FABPC gene, was isolated from a Sj lambda ZAPII cDNA expression library immunoscreened with hyperimmune rabbit serum (HRS) raised against soluble adult Sj proteins. The complete ORF (open reading frame) of Sj-FABPC encodes a protein of 132 amino acids (aa) of 14.7 kDa. The aa sequence of Sj-FABPC exhibits 91% identity to a FABP of S. mansoni (Sm14) and 45% identity to a FABP of Fasciola hepatica (Fh15), putative vaccine candidates for schistosomiasis. Sj-FABPC was subcloned into the QIAexpress vector, pQE-10, and subsequently expressed in Escherichia coli. The re-Sj-FABPC, purified under non-denaturing conditions, was recognized by sera from patients with acute and chronic schistosomiasis japonica. The purified re-Sj-FABPC was also shown to bind to palmitic acid with high affinity. The functional expression of Sj-FABPC will facilitate studies on re-Sj-FABPC to assess its potential as a drug and/or vaccine candidate.

Amino Acid Sequence↗
PubMed1994

Purification of a recombinant Schistosoma japonicum antigen homologous to the 22-kDa membrane-associated antigen of S. mansoni, a putative vaccine candidate against schistosomiasis.

We describe the cDNA cloning, overproduction and purification of a 22.6-kDa antigen from the human blood fluke Schistosoma japonicum. A 777-bp cDNA (C32) was isolated from a S. japonicum lambda ZAPII cDNA expression library immuno-screened with hyperimmune rabbit serum (HRS) raised against soluble adult S. japonicum proteins. The open reading frame of C32 encodes a protein of 191 amino acids (aa) which exhibits 71% identity to a 22.6-kDa membrane-associated antigen of S. mansoni, a putative vaccine candidate for schistosomiasis. We have identified a sequence motif known as an EF-hand calcium-binding domain in both the S. japonicum and S. mansoni aa sequences, suggesting that the 22.6-kDa antigens are able to bind Ca2+. Further, we have, for the first time, obtained the 22.6-kDa antigen in purified, non-denatured, recombinant form, and in sufficient quantity to assess the protective value of the molecule in vaccination/challenge experiments. This was achieved by synthesizing the schistosome antigen with a short polyhistidine tag fused to the N-terminus which was then used for subsequent affinity purification. The recombinant protein was purified under non-denaturing conditions using nickel-chelate affinity chromatography.

Amino Acid Sequence↗