The Effect of Alpha-mating Factor Secretion Signal Mutations on Recombinant Protein Expression in Pichia pastoris
Poster Number
37
Format
Poster Presentation
Faculty Mentor Name
Joan Lin-Cereghino
Faculty Mentor Department
Biological Sciences
Additional Faculty Mentor Name
Geoffrey Lin-Cereghino
Abstract/Artist Statement
Pichia pastoris is a species of methylotrophic yeast used as an expression system to produce and secrete foreign proteins. Secretion of such heterologous proteins requires fusion of alphamating factor secretion signal (Mat-alpha) to the N-terminus of the protein construct. The secretion signal acts as an "address label" that helps direct the protein to be secreted out of the cell. Here, we describe how the removal and addition of specific amino acids in the 85 amino acid Mat-alpha secretion signal affects the secretion of horseradish peroxidase (HRP), which was used as a reporter. Computer modeling of the Mat-alpha secondary structure created in the Lin-Cereghino lab as well as bioinformatics resources were used to create mutant Mat-alpha sequences in order to comprehend how the signal functions. The results allowed us to not only understand the potential contribution of each segment, but also ponder possibilities to achieve higher levels of secretion. Several mutant signals were found to provide significantly greater secretion levels of HRP than the wild type Mat-alpha, indicating that the targeted amino acids could play an essential role in the export pathway.
Location
DeRosa University Center, Ballroom
Start Date
26-4-2014 2:00 PM
End Date
26-4-2014 4:00 PM
The Effect of Alpha-mating Factor Secretion Signal Mutations on Recombinant Protein Expression in Pichia pastoris
DeRosa University Center, Ballroom
Pichia pastoris is a species of methylotrophic yeast used as an expression system to produce and secrete foreign proteins. Secretion of such heterologous proteins requires fusion of alphamating factor secretion signal (Mat-alpha) to the N-terminus of the protein construct. The secretion signal acts as an "address label" that helps direct the protein to be secreted out of the cell. Here, we describe how the removal and addition of specific amino acids in the 85 amino acid Mat-alpha secretion signal affects the secretion of horseradish peroxidase (HRP), which was used as a reporter. Computer modeling of the Mat-alpha secondary structure created in the Lin-Cereghino lab as well as bioinformatics resources were used to create mutant Mat-alpha sequences in order to comprehend how the signal functions. The results allowed us to not only understand the potential contribution of each segment, but also ponder possibilities to achieve higher levels of secretion. Several mutant signals were found to provide significantly greater secretion levels of HRP than the wild type Mat-alpha, indicating that the targeted amino acids could play an essential role in the export pathway.