TY - JOUR
T1 - Molecular cloning and characterization of a C-type lectin from Ancylostoma ceylanicum
T2 - Evidence for a role in hookworm reproductive physiology
AU - Brown, Allison C.
AU - Harrison, Lisa M.
AU - Kapulkin, Wadim
AU - Jones, Brian F.
AU - Sinha, Anindita
AU - Savage, Amy
AU - Villalon, Nicholas
AU - Cappello, Michael
N1 - Funding Information:
The authors would like to thank Gerhard Schad and Alan Scott for thoughtful interpretation of the immunohistochemistry data, and Richard Bungiro for advice during the course of this work. This work was supported by grant AI47929 from the National Institutes of Health (MC). MC is a recipient of a Hellman Family Fellowship from the Office of the President of Yale University.
PY - 2007/2
Y1 - 2007/2
N2 - Lectins comprise a family of related proteins that mediate essential cell functions through binding to carbohydrates. Within this protein family, C-type lectins are defined by the requirement of calcium for optimal biologic activity. Using reverse transcription PCR, a cDNA corresponding to a putative C-type lectin has been amplified from the hookworm parasite Ancylostoma ceylanicum. The 550 nucleotide open reading frame of the A. ceylanicum C-type Lectin-1 (AceCTL-1) cDNA corresponds to a 167 amino acid mature protein (18,706 Da) preceded by a 17 amino acid secretory signal sequence. The recombinant protein (rAceCTL-1) was expressed in Drosophila S2 cells and purified using a combination of affinity chromatography and reverse phase HPLC. Using in vitro carbohydrate binding studies, it was determined that rAceCTL-1 binds N-acetyl-d-glucosamine, a common component of eukaryotic egg cell membranes. Using a polyclonal IgG raised against the recombinant protein, the native AceCTL-1 was identified in sperm and soluble protein extracts of adult male A. ceylanicum by immunoblot. Probing of adult hookworm sections with the polyclonal IgG demonstrated localization to the testes in males, as well as the spermatheca and developing embryos in females, consistent with its role as a sperm protein. Together, these data strongly suggest that AceCTL-1 is a male gender-specific C-type lectin with a function in hookworm reproductive physiology.
AB - Lectins comprise a family of related proteins that mediate essential cell functions through binding to carbohydrates. Within this protein family, C-type lectins are defined by the requirement of calcium for optimal biologic activity. Using reverse transcription PCR, a cDNA corresponding to a putative C-type lectin has been amplified from the hookworm parasite Ancylostoma ceylanicum. The 550 nucleotide open reading frame of the A. ceylanicum C-type Lectin-1 (AceCTL-1) cDNA corresponds to a 167 amino acid mature protein (18,706 Da) preceded by a 17 amino acid secretory signal sequence. The recombinant protein (rAceCTL-1) was expressed in Drosophila S2 cells and purified using a combination of affinity chromatography and reverse phase HPLC. Using in vitro carbohydrate binding studies, it was determined that rAceCTL-1 binds N-acetyl-d-glucosamine, a common component of eukaryotic egg cell membranes. Using a polyclonal IgG raised against the recombinant protein, the native AceCTL-1 was identified in sperm and soluble protein extracts of adult male A. ceylanicum by immunoblot. Probing of adult hookworm sections with the polyclonal IgG demonstrated localization to the testes in males, as well as the spermatheca and developing embryos in females, consistent with its role as a sperm protein. Together, these data strongly suggest that AceCTL-1 is a male gender-specific C-type lectin with a function in hookworm reproductive physiology.
KW - Ancylostoma ceylanicum
KW - C-type lectin
KW - Hookworm
KW - Nematode
KW - Sperm
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U2 - 10.1016/j.molbiopara.2006.10.017
DO - 10.1016/j.molbiopara.2006.10.017
M3 - Article
C2 - 17129620
AN - SCOPUS:33846003857
SN - 0166-6851
VL - 151
SP - 141
EP - 147
JO - Molecular and Biochemical Parasitology
JF - Molecular and Biochemical Parasitology
IS - 2
ER -