Periplasmic expression and recovery of human interferon gamma in Escherichia coli.
A synthetic human interferon gamma (hIFN-gamma) gene was fused to SP1 and SP3, two Sec-dependent artificial signal peptides to transport the hIFN-gamma to the periplasm of Escherichia coli BL21-SI. The processing efficiency of both SP1-hIFN-gamma and SP3-hIFN-gamma was dependent on the culture medium as well as the post-induction temperature. Both precursors were processed completely when cells were cultured using minimal medium and a post-induction temperature of 32.5 degrees C, and only the processed hIFN-gamma was detected. The SP3 signal peptide was more efficient than SP1 for the secretion of hIFN-gamma. Sixty percent of the total hIFN-gamma was secreted to the periplasm using the SP3 signal peptide and a post-induction temperature of 20 degrees C. Using Tris-sucrose-dithiothreitol (TSD) hypertonic buffer, the periplasmic soluble hINF-gamma was recovered with a purity of 85%.
Balderas Hernández
Victor E
VE
División de Biología Molecular, Instituto Potosino de Investigación Científica y Tecnológica, Camino a la Presa San José 2055, Col. Lomas 4a. Sección, C.P. 78216 San Luis Potosí, SLP, Mexico.
Paz Maldonado
Luz M T
LM
Medina Rivero
Emilio
E
Barba de la Rosa
Ana P
AP
Jiménez-Bremont
Juan F
JF
Ordoñez Acevedo
Leandro G
LG
De León Rodríguez
Antonio
A
D000595:Amino Acid Sequence; D001483:Base Sequence; D004926:Escherichia coli / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D019897:Periplasm / Q000378:metabolism*; D021382:Protein Sorting Signals / Q000235:genetics; D013696:Temperature
D021382:Protein Sorting Signals; D007371:Interferon-gamma