Haemonchus contortus: evaluation of two signal sequence trapping systems for detection of secreted molecules.
Given that signal sequences between secreted proteins of different species can be interchanged, it is reasonable to expect that both mammalian and yeast signal sequence trapping (SST) systems would secrete Haemonchus contortus proteins with similar efficiency and quality. To determine if H. contortus cDNAs that contain a signal sequence could re-establish secretion of a reporter protein, mammalian and yeast SST vectors were designed, 10 H. contortus genes selected, and their respective cDNAs cloned into these two SST vectors. The selected molecules included genes known to code for excretory/secretory or membrane-bound proteins as potential test 'positives', and genes known to code for non-secreted proteins as test 'negatives'. While differentiation between secretion and non-secretion was evident in both systems, the results indicated greater efficiency was achieved when the mammalian system was used. Therefore, mammalian SST using COS cells would be a more useful tool to screen H. contortus cDNA libraries for potential secreted and type-1 integral membrane proteins than yeast SST.
Donald
Debbie
D
Department of Primary Industries Victoria, 475 Mickleham Road, Attwood, Vic. 3049, Australia.
Chen
Yaping
Y
Hartman
Dadna
D
Zawadzki
Jody L
JL
D000681:Amylases / Q000235:genetics; D000818:Animals; D018826:CD13 Antigens / Q000235:genetics / Q000502:physiology; D019556:COS Cells; D002403:Cathepsins / Q000235:genetics / Q000502:physiology; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D005822:Genetic Vectors; D006189:Haemonchus / Q000235:genetics* / Q000378:metabolism*; D015801:Helminth Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014162:Transfection
D015801:Helminth Proteins; D008565:Membrane Proteins; D021382:Protein Sorting Signals; D000681:Amylases; D002403:Cathepsins; D018826:CD13 Antigens; C080908:H11 protein, Haemonchus contortus