Electrophoretic Extraction and Proteomic Characterization of Proteins Buried in Marine Sediments

Abstract

Abstract:Proteins are the largest defined molecular component of marine organic nitrogen, and hydrolysable amino acids, the building blocks of proteins, are important components of particulate nitrogen in marine sediments. In oceanic systems, the largest contributors are phytoplankton proteins, which have been tracked from newly produced bloom material through the water column to surface sediments in the Bering Sea, but it is not known if proteins buried deeper in sediment systems can be identified with confidence. Electrophoretic gel protein extraction methods followed by proteomic mass spectrometry and database searching were used as the methodology to identify buried phytoplankton proteins in sediments from the 8-10 cm section of a Bering Sea sediment core. More peptides and proteins were identified using an SDS-PAGE tube gel than a standard 1D flat gel or digesting the sediment directly with trypsin. The majority of proteins identified correlated to the marine diatom, Thalassiosira pseudonana, rather than bacterial protein sequences, indicating an algal source not only dominates the input, but also the preserved protein fraction. Abundant RuBisCO and fucoxanthin chlorophyll a/c binding proteins were identified, supporting algal sources of these proteins and reinforcing the proposed mechanisms that might protect proteins for long time periods. Some preserved peptides were identified in OPEN ACCESSChromatography 2014, 1 177 unexpected gel molecular weight ranges, indicating that some structural changes or charge alteration influenced the mobility of these products during electrophoresis isolation. Identifying buried photosystem proteins suggests that algal particulate matter is a significant fraction of the preserved organic carbon and nitrogen pools in marine sediments.

Diversity of Cultivable Protease-Producing Bacteria in Laizhou Bay Sediments, Bohai Sea, China

“…These processes are mainly carried out by bacteria that produce degradation enzymes (Brunnegård et al, 2004 ; Hunter et al, 2006 ). Since proteins comprise the main component of biomass of marine organisms (Thamdrup and Dalsgaard, 2008 ; Lloyd et al, 2013 ; Moore et al, 2014 ), protease-producing bacteria are considered as the main degraders of organic nitrogen in the marine environment (Chen et al, 2003 ; Zhao et al, 2012 ). These protease-producing bacteria usually secrete extracellular proteases that degrade protein materials, for example, by hydrolyzing the OrgN into peptides and amino acids, with the latter easily taken up by bacteria for subsequent catabolism (Zhao et al, 2012 ).…”

Diversity of cultivable protease-producing bacteria in sediments of Jiaozhou Bay, China

“…Microbial enzymatic hydrolysis of these nitrogenous macromolecules is essential for the mineralization of sedimentary organic nitrogen (SON) and the benthic nutrients recycling (Talbot and Bianchi, 1997 ; Fabiano and Danovaro, 1998 ; Herbert, 1999 ; Patel et al, 2001 ; Thamdrup and Dalsgaard, 2008 ; Arnosti, 2011 ; Arnosti et al, 2014 ). As proteins constitute large fractions of marine organic matters (Wakeham et al, 1997 ; Thamdrup and Dalsgaard, 2008 ; Lloyd et al, 2013 ; Moore et al, 2014 ), protease-producing bacteria are recognized as key players in the microbial degradation of SON (Herbert, 1999 ; Zhao et al, 2008 , 2012 ; Chen et al, 2009 ; Zhou et al, 2009 ). They secrete extracellular proteases to hydrolyze complex proteinaceous substances into small peptides and amino acids suitable for cellular uptake (Zhao et al, 2008 , 2012 ), initiating the mineralization of SON and driving the nitrogen cycle in marine ecosystem.…”

Progress and Challenges in Ocean Metaproteomics and Proposed Best Practices for Data Sharing

“…25,32 Moreover, the presence of biogenic soft and hard parts, including mucilage, calcium carbonate, and siliceous components, as well as mineral phases, can complicate chemical separation of proteins and impair protein extraction efficiencies and require development of matrix-specific extraction protocols. 6,12,17,22,33 Mass Spectrometry Analyses…”