Title

The All-Powerful Universal Secretor: Does it Really Exist?

Poster Number

25

Format

Poster Presentation

Abstract/Artist Statement

Heterologous expression is the practice of using microorganisms to synthesize proteins they usually do not make. It has been useful in industry to make mass quantities of vital human proteins such as insulin. One microorganism, Pichia pastoris, is a popular strain of yeast selected by many academic and industrial laboratories for expression of these heterologous proteins. To date, over 800 proteins have been successfully expressed using P. pastoris’ powerful secretory machinery. Despite this fact, there are some proteins that P. pastoris is unable to efficiently express in significant quantities. Preceding this project, mutant super-secretor strains of P. pastoris were created and found to secrete substantial amounts of β-galactosidase; their disrupted genes were identified using BLAST sequence analysis, and these genes were isolated and cloned. Our current objective is to further investigate expression levels of two proteins HRP and SLPI in super-secretor mutants LL1, AH1 and AH2, compared to the expression levels observed in wild- type strains. Our experimental data suggest that the amounts of HRP and SLPI secreted by LL1, AH1 and AH2 slightly increased. The next steps of this study will include the development of knockouts to further examine the intricacy of Pichia’s secretory mechanisms and pinpoint the key to utilizing it to its full potential—becoming a more universal secretor.

Location

DeRosa University Center, Ballroom B

Start Date

1-5-2010 1:00 PM

End Date

1-5-2010 3:00 PM

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May 1st, 1:00 PM May 1st, 3:00 PM

The All-Powerful Universal Secretor: Does it Really Exist?

DeRosa University Center, Ballroom B

Heterologous expression is the practice of using microorganisms to synthesize proteins they usually do not make. It has been useful in industry to make mass quantities of vital human proteins such as insulin. One microorganism, Pichia pastoris, is a popular strain of yeast selected by many academic and industrial laboratories for expression of these heterologous proteins. To date, over 800 proteins have been successfully expressed using P. pastoris’ powerful secretory machinery. Despite this fact, there are some proteins that P. pastoris is unable to efficiently express in significant quantities. Preceding this project, mutant super-secretor strains of P. pastoris were created and found to secrete substantial amounts of β-galactosidase; their disrupted genes were identified using BLAST sequence analysis, and these genes were isolated and cloned. Our current objective is to further investigate expression levels of two proteins HRP and SLPI in super-secretor mutants LL1, AH1 and AH2, compared to the expression levels observed in wild- type strains. Our experimental data suggest that the amounts of HRP and SLPI secreted by LL1, AH1 and AH2 slightly increased. The next steps of this study will include the development of knockouts to further examine the intricacy of Pichia’s secretory mechanisms and pinpoint the key to utilizing it to its full potential—becoming a more universal secretor.