Date Approved

2017

Degree Type

Open Access Senior Honors Thesis

Department or School

Biology

First Advisor

Aaron Liepman, PhD

Second Advisor

Kristin Judd, PhD

Abstract

Mannans are a type of hemicellulosic plant cell wall polysaccharide that offer structural support and serve as energy reserves for developing plants. Coffee seeds are rich in mannans, and mannan content of coffee seeds impacts brew quality. Mannan backbones are synthesized by CSLA proteins. This study aims to characterize a CSLA enzyme from Coffea canephora (mannan synthase 1; CcMANS1) that has been implicated in the biosynthesis of backbones of coffee seed mannans. This enzyme has not been biochemically characterized and characterizing its activity will help contribute to an understanding of the differences between CSLA proteins that dictate the composition of carbohydrate products. Recombinant CcMANS1 was produced using transgenic Pichia pastoris and Saccharomyces cerevisiae. Despite the use of multiple assays, including biochemical assays, fluorophore assisted carbohydrate electrophoresis, and mass spectroscopy, it was not possible to confirm mannan synthase activity of recombinant CcMANS1. Future studies that characterize additional putative mannan synthases or that focus on identifying the cause(s) of inactivity of CcMANS1 may prove beneficial.

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