Proteins constitute one of the largest fractions of organic material in secondary effluent from domestic wastewater treatment plants, and are a significant source of organic N released into receiving waters. Wastewater effluents from two wastewater treatment plants in Western Massachusetts were analyzed for protein content and activity. Effluents were concentrated using ammonium sulfate precipitation, and the component proteins subsequently separated using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and zymogramic electrophoresis. The results showed that some protein bands in primary effluent are also present in the secondary effluent, indicating the recalcitrance of some sewage proteins. These proteins are released into the Connecticut River. The presence of these same bands in secondary effluent that was filtered through a 0.45µm filter is an indication that a filtration step would not prevent the release of these particular proteins. Other bands that are visible in the secondary effluent are not found in the primary effluent, thus those proteins are produced in the activated sludge process. Zymogram electrophoresis of concentrated and raw effluents showed that some of the proteins in both primary and secondary effluent are proteolytic enzymes. The parameters of total protein, inorganic nitrogen, organic nitrogen, COD, and TSS were also measured from primary and pre-chlorinated secondary effluents from each plant. It is thought that any proteins in secondary effluent are not biologically available given that they have survived the treatment processes in the plants; however, this has not been shown. This initial characterization of proteins is the first step in determining the impact of proteins in wastewater effluent on receiving waters. Future work will focus on identifying individual proteins and building a protein dataset that will enable tracking of effluent proteins in the environment.