Rubisco Case Study

titel

achterkant

Voorwoord

Samenvatting

Table of Contents
List of abbreviations 1
1. Introduction 2
1.1 Biobased products 2
1.2 RuBisCO 3
1.3 Isochrysis galbana 4
1.4 Tetraselmis sp 4
2. Methods 5
2.1 Size Exclusion Chromatography 5
2.2 SDS-PAGE 6
2.3 Bradford protein assay 7
2.4 Ion Exchange Chromatography 7
2.5 Soxhlet extraction method 8
2.6 Kjeldahl method 8
3. Materials 9
3.1 Size Exclusion Chromatography 9
3.2 SDS-PAGE 9
3.3 Bradford protein assay 9
3.4 Ion Exchange Chromatography 10
3.5 Soxhlet extraction method 10
3.6 Kjeldahl method 10
3.7 Enzymatic digestion 10
3.8 Used strains 11
3.9 General protein extraction method 11
4. Results 12

RuBisCO has a catalytic rate of a few CO2 molecules per second at 25 °C 19. Besides it has a low affinity for atmospheric CO2 and uses O2 as an alternative substrate. For these reasons RuBisCO is an inefficient enzyme as the initial CO2-fixing enzyme for photosynthesis. Therefore, plants and algae must contribute a major part of their nitrogen to RuBisCO. The total soluble protein content inside the plants can comprise up to 50%20.
Because RuBisCO is so abundantly present in plants it was assumed that the same applies to microalgae. In 1991, J.A. Raven compiled a dataset for RuBisCO in microalgae as the fraction of RuBisCO to total protein by mass, showing values between 2% and 23%. To determine these percentages however, an indirect measurement was performed where RuBisCO is converted on a total protein basis assuming the chlorophyll: total protein mass ratio is 0.0421.
Therefore, Losh et al. conducted experiments where RuBisCO was measured with Quantitative Western blots using an antibody which binds to a conserved region of the large subunit of RuBisCO. They concluded that RuBisCO represented < 6% of total protein in eight species of microalgae. Furthermore, they concluded that unlike in plants, RuBisCO does not account for a major fraction of cellular nitrogen in