Abstract:
The green alga Botryococcus braunii was degraded for 201 days under oxic conditions in a flow-through system to test whether the macromolecular algaenan surrounding cells could protect proteins from rapid degradation. Protein loss was ~8x slower for B. braunii than for other previously studied phytoplankton, with base extraction consistently removing only a small fraction (<35%) of the total proteinaceous material compared to that seen by acid hydrolysis. Size-exclusion chromatography coupled to evaporative laser-light scattering detection and fluorescence spectroscopy detected proteinaceous materials from 1.7 x 104 to > 1.5 x 106 relative molecular mass (Mr) preserved in degraded material. A shift to higher Mr material during B. braunii decay and response of protein aggregates to denaturing agents identified hydrophobic and hydrogen-bond interactions as important stabilizing forces for protein preservation. Two-dimensional electrophoresis of proteins extractable from detrital material indicated the predominant presence of modified proteins. Susceptibility of the detrital organic matter to cleavage with proteolytic enzymes indicated the retention of peptide bonds, while incomplete cleavage of aggregates may be due to associations of proteins with algaenans. Using an antibody probe for RuBisCo, the large subunit was not retained in its original form (Mr 55,000), but additional cross-linked or other aggregated products (> Mr 73,000) plus a cleavage product (Mr 24,000) were observed during the decay sequence. In the 201-day-old B. braunii detritus, the large contribution of nonpolar > 2-kDa-hydrolyzable amino acids to bulk nitrogen (~50%) suggests that encapsulation and hydrophobic associations protect a fraction of protein from bacterial degradation, and allow for preservation of their products seen as amino acids in the Pula kerogen.
