Identification, isolation, and molecular cloning of a hookworm protease: an approach towards a defined vaccine for ancylostomiasis
Thesis/Dissertation
·
OSTI ID:6591698
The hookworm Ancylostoma caninum was shown to release in vitro a 37 kDa protease that catalyzed the hydrolysis of fibrinogen, plasminogen, and elastin. The enzyme was purified from parasite extracts by ion-exchange chromatography, followed by gel filtration and hydrophobic interaction chromatography. An amino-terminal sequence was determined. When assayed with radiolabeled fibrin as substrate, the enzyme displayed optimal activity at pH 9-11; it was inactivated by dialysis against ethylenediamine tetraacetic acid. Antiserum raised against the protease in rabbits cross-reacted on western blots with soluble antigen from the infective larval stage of the parasite. A cDNA library from hookworm mRNA was constructed in the expression vector bacteriophage lambdagtll. A positive clone was identified with the rabbit antiserum that was shown to contain an 800-bp insert. The insert was mapped, subcloned into M13, and sequenced, revealing an open reading frame of 789 nucleotides corresponding to 263 amino acids.
- Research Organization:
- Rockefeller Univ., New York (USA)
- OSTI ID:
- 6591698
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
550201* -- Biochemistry-- Tracer Techniques
550901 -- Pathology-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACID SEQUENCE
CHEMICAL COMPOSITION
CHROMATOGRAPHY
CLONING
COAGULANTS
DISEASES
DNA-CLONING
DRUGS
ENZYMES
FIBRIN
FRACTIONATION
HEMATOLOGIC AGENTS
HEMOSTATICS
HYDROLASES
INFECTIOUS DISEASES
ION EXCHANGE CHROMATOGRAPHY
MESSENGER-RNA
MOLECULAR STRUCTURE
NUCLEIC ACIDS
ORGANIC COMPOUNDS
PARASITES
PARASITIC DISEASES
PATHOGENESIS
PEPTIDE HYDROLASES
PROTEINS
RADIOASSAY
RNA
SCLEROPROTEINS
SEPARATION PROCESSES
VACCINES
550901 -- Pathology-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACID SEQUENCE
CHEMICAL COMPOSITION
CHROMATOGRAPHY
CLONING
COAGULANTS
DISEASES
DNA-CLONING
DRUGS
ENZYMES
FIBRIN
FRACTIONATION
HEMATOLOGIC AGENTS
HEMOSTATICS
HYDROLASES
INFECTIOUS DISEASES
ION EXCHANGE CHROMATOGRAPHY
MESSENGER-RNA
MOLECULAR STRUCTURE
NUCLEIC ACIDS
ORGANIC COMPOUNDS
PARASITES
PARASITIC DISEASES
PATHOGENESIS
PEPTIDE HYDROLASES
PROTEINS
RADIOASSAY
RNA
SCLEROPROTEINS
SEPARATION PROCESSES
VACCINES