Adam Lepczyński

Occurrence of aspartyl proteases in brine after herring marinating

Herrings are marinated in a brine consisting of salt and acetic acid. During marinating, various nitrogen fractions diffuse from fish flesh to the brine, causing significant nutritional quality losses of the raw material. In this study, it has been demonstrated for the first time that proteases diffuse from the fish to the marinating brine. Using ammonium sulphate precipitation and affinity chromatography on pepstatin-A agarose bed the aspartyl proteases were purified and concentrated over 2600-fold from a marinating brine. Pepstatin-A completely inhibited the activity of the purified preparation. The preparation was active against fluorogenic substrates specific for cathepsin D and E and inactive against substrates specific for cysteine cathepsins. Depending on incubation time, the preparation showed pH-optimum at 2.0 or 4.5. The 2D SDS-PAGE separation demonstrated the presence of a few proteins with molecular weights and pI values typical of cathepsin D, E and pepsin.

Immunohistochemical identification of aquaporin 2 in the kidneys of young beef cattle

Abstract Aquaporin 2 (AQP2) is a small, integral tetrameric plasma membrane protein that is expressed in mammalian kidneys. The specific constitution of this protein and its selective permeability to water means that AQP2 plays an important role in hypertonic urine production. Immunolocalization of AQP2 has been studied in humans, monkeys, sheep, dogs, rabbits, rats, mice and adult cattle. We analyzed the expression of AQP2 in kidneys of 7-month-old Polish-Friesian var. black and white male calves. AQP2 was localized in the principal cells of collecting ducts in medullary rays penetrating the renal cortex and in the collecting ducts of renal medulla. AQP2 was expressed most strongly in the apical plasma membrane, but expression was observed also in the intracellular vesicles and basolateral plasma membrane. Our study provides new information concerning the immunolocalization of AQP2 in calf kidneys.

Human sperm proteins identified by 2-dimensional electrophoresis and mass spectrometry and their relevance to a transcriptomic analysis

The aim of this study was to identify and analyse human sperm proteins from normozoospermic men using 2-dimensional electrophoresis (2-DE) and mass spectrometry (MS). We identified 73 different sperm proteins, including two less characterized human sperm proteins, Annexin A7 (ANXA7) and c14orf105. Bioinformatic analysis of detected sperm proteins revealed new carbohydrate and lipid metabolic pathways, which supply energy to motile sperm. A comparison of our data with available mRNA microarray data from the human testis allows for validation of identified sperm proteins and aids in the recognition of their physiological pathways.

Potential plasma biomarkers of bladder canceridentified by proteomic analysis: A pilot study

BACKGROUND Bladder cancer diagnosis and surveillance includes cystoscopy and cytology. New methods for the detection of bladder cancer are needed, because cystoscopy is invasive and expensive, and because urine cytology is not sensitive enough. OBJECTIVES The aim of the study was to select potential plasma protein markers for bladder cancer which could be useful in developing a specific laboratory test to improve diagnosis and to establish treatment strategies in order to prevent the recurrence of the disease. MATERIAL AND METHODS Plasma proteome maps were prepared based on 2-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), combined with image gel analysis and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry of plasma samples from patients with urothelial bladder cancer, and they were compared to normal samples. RESULTS The analyses of bladder cancer plasma samples allowed us to distinguish 3 groups of proteins whose relative abundance differed from that in normal samples. The 1st one comprised modified forms of plasma transferrin, fibrinogen gamma and complement C3b, which were absent in normal plasma. The 2nd group comprised haptoglobin, alpha-2-macroglobulin, vitamin D-binding protein, and pigment epithelium-derived factor, which occurred in the cancerous samples in large quantities. The 3rd group consisted of 3 molecular forms of immunoglobulin M (IgM), the relative abundance of which was significantly lower in the cancerous plasma samples. CONCLUSIONS The data indicated potential plasma biomarkers associated with inflammation, immunity and coagulation processes accompanying bladder cancer. They could be used for the development of a laboratory test(s) useful in clinical practice.