Nitrogen (N) is a major nutrient needed for plant growth. Around 40% of N is considered unavailable for plant use in the form of proteins. Proteins can be broken down into constituents available for plants by extracellular proteases released by microorganisms. Therefore, understanding soil proteases and their role in organic matter breakdown could provide a mechanism for increasing organic N acquisition that would improve N supply in soil leading to increased plant productivity. However, there are ongoing discussions over the lack of standardisation of assays used to measure soil protein and protease activity. The overall aims of the thesis were to a) critically evaluate the methods used to extract proteins from soil and measure soil protease activity and b) determine the factors that influence protein breakdown in the soil-plant-microorganism system. First, I critically evaluated methods to extract proteins from soil by comparing the ability of common extractants to recover soluble proteins from three soil types. I found that the dependence of protein recovery on both extractant and soil type prevents direct comparison of studies using different recovery methods, particularly if no extraction controls are used. Secondly, I investigated how topsoil and subsoil properties affect protein breakdown along a grassland altitude and primary productivity gradient that contained a range of soil types. I concluded that protein breakdown was not regulated by a small number of factors but a wide range of interacting factors which were site specific. Furthermore, I suggested that differences in soil N cycling and the generation of ammonium are more related to the rate of protein supply rather than limitations in protease activity and protein turnover. I then determined whether plants actively secrete proteases to enhance the breakdown of soil protein or are they functionally reliant on soil microorganisms to undertake this role? The results indicated that plant uptake of organic N is only functionally significant when soil protein is in direct contact with root surfaces. The lack of protease upregulation under N deficiency suggests that root protease activity is unrelated to enhanced soil N capture. Next, I determined the spatial distribution of protease activity in the rhizosphere of barley (Hordeum vulgare L.) using in situ zymography. I analysed the effect of root hairs and soluble protein addition on rhizosphere protease activity. The results showed protease activity was highest in the barley genotype with root hairs and with protein addition suggesting that plants with root hairs have a greater advantage in accessing protein hotspots in the soil indirectly via microbial-derived proteases. Lastly, I conducted a systematic review and meta-analysis of the common methods used to measure soil protease activity globally. I collected data on environmental and methodological factors to determine the variation of protease activity in soil. From this, I found soil protease activity to vary widely due to study-biases and observed a lack of reporting of key assay conditions by studies. Together, this research provides a more detailed understanding of protein mineralisation and protease activity in soil. Going forward, comprehensive reporting of enzyme assay conditions is essential to increase the accuracy and reliability of interpreting soil protease activity dynamics in the soil-plant-microorganism system.