Sensitive method to identify and characterize proteinases in situ after SDS-PAGE.
AuthorsWilliams J, McGrath WJ, Mangel WF
JournalBiotechniques
PubMed ID11084874
'Cells and body fluids contain numerous, different proteinases; to identify and characterize them are both important and difficult tasks. Especially difficult to identify and characterize are highly specific proteinases. Here, we present an extremely sensitive and quantitative method to characterize proteinases fractionated by SDS-PAGE that cleave specific rhodamine-based fluorogenic substrates. ... More
Activation of trypsinogen in large endocytic vacuoles of pancreatic acinar cells.
'The intracellular activation of trypsinogen, which is both pH- and calcium-dependent, is an important early step in the development of acute pancreatitis. The cellular compartment in which trypsinogen activation occurs currently is unknown. We therefore investigated the site of intracellular trypsinogen activation by using an established cellular model of acute ... More
activated polyamine catabolism in acute pancreatitis: alpha-methylated polyamine analogues prevent trypsinogen activation and pancreatitis-associated mortality.
AuthorsHyvönen MT, Herzig KH, Sinervirta R, Albrecht E, Nordback I, Sand J, Keinänen TA, Vepsäläinen J, Grigorenko N, Khomutov AR, Krüger B, Jänne J, Alhonen L
JournalAm J Pathol
PubMed ID16400014
'Polyamines are essential for normal cellular growth and function. Activation of polyamine catabolism in transgenic rats overexpressing spermidine/spermine N(1)-acetyltransferase, the key enzyme in polyamine catabolism, results in severe acute pancreatitis. Here, we investigated the role of polyamine catabolism in pancreatitis and studied the effect of polyamine analogues on the outcome ... More
Direct detection of premature protease activation in living pancreatic acinar cells.
AuthorsKrüger B, Lerch MM, Tessenow W
JournalLab Invest
PubMed ID9645767
A microspectrofluorometric study of porphyrin-photosensitized single living cells--I. Membrane alterations.
Rhodamines 110 and 123, and rhodamine 110 linked via peptide bonds to Arg, Cbz-Arg and Cbz-Ile-Pr-Arg interact with free base porphyrins or cytochrome C. Rhodamines 110 and 123 essentially form 1:1 complexes while the other derivatives form 2:1 complexes. The possible biological implications of these results are discussed. ... More
A homogeneous, nonradioactive high-throughput fluorogenic protein kinase assay.
AuthorsKupcho K, Somberg R, Bulleit B, Goueli SA
JournalAnal Biochem
PubMed ID12758259
Protein kinases play an important role in many cellular processes and mediate cellular responses to a variety of extracellular stimuli. They have been identified by many pharmaceuticals as valid targets for drug discovery. Because of the large number of protein kinases, and the large number of compounds to be screened, ... More
A homogeneous, nonradioactive high-throughput fluorogenic protein phosphatase assay.
AuthorsKupcho K, Hsiao K, Bulleit B, Goueli SA
JournalJ Biomol Screen
PubMed ID15140384
Protein phosphatases are critical components in cellular regulation; they do not only act as antioncogenes by antagonizing protein kinases, but they also play a positive regulatory role in a variety of cellular processes that require dephosphorylation. Thus, assessing the function of these enzymes necessitates the need for a robust, sensitive ... More
Calcium-dependent enzyme activation and vacuole formation in the apical granular region of pancreatic acinar cells.
The pancreatic acinar cell produces powerful digestive enzymes packaged in zymogen granules in the apical pole. Ca(2+) signals elicited by acetylcholine or cholecystokinin (CCK) initiate enzyme secretion by exocytosis through the apical membrane. Intracellular enzyme activation is normally kept to a minimum, but in the often-fatal human disease acute pancreatitis, ... More
Trypsin activity is not involved in premature, intrapancreatic trypsinogen activation.
AuthorsHalangk W, Krüger B, Ruthenbürger M, Stürzebecher J, Albrecht E, Lippert H, Lerch MM
JournalAm J Physiol Gastrointest Liver Physiol
PubMed ID11804859
A premature and intracellular activation of digestive zymogens is thought to be responsible for the onset of pancreatitis. Because trypsin has a critical role in initiating the activation cascade of digestive enzymes in the gut, it has been assumed that trypsin also initiates intracellular zymogen activation in the pancreas. We ... More