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Biomedical subjects

M Finken

Publications and source records attributed to M Finken.

6 recordsLinked to original sources
PubMed1994

Characterization of the complete protein disulfide isomerase gene of Schistosoma mansoni and identification of the tissues of its expression.

cDNA and genomic clones of Schistosoma mansoni containing the complete sequence of a homolog of protein disulfide isomerase have been identified. The protein disulfide isomerase gene in schistosomes is a single copy gene having a genomic structure that is very similar to that of man. The C-terminus of the deduced protein is KDEL which in mammals functions as a signal sequence for retention of luminal proteins in the endoplasmic reticulum. Immunohistology and in situ hybridization identify the gastrodermis of the gut, the wall cells of the protonephridia, and the sustentacular cells of the testes to be the major tissues of protein disulfide isomerase gene expression. The protein disulfide isomerase of schistosomes, produced in an expression vector in Escherichia coli, catalyzes disulfide/sulfhydryl isomerization in vitro.

Amino Acid Sequence↗
PubMed1993

Molecular basis of streptomycin resistance in Mycobacterium tuberculosis: alterations of the ribosomal protein S12 gene and point mutations within a functional 16S ribosomal RNA pseudoknot.

Multidrug-resistant strains of Mycobacterium tuberculosis have resulted in several recent outbreaks. Recognition of drug resistance is important both for treatment and to prevent further transmission. Here we use molecular biology techniques to study the basis of streptomycin resistance in single and multidrug-resistant M. tuberculosis. We demonstrate that streptomycin resistance is associated with mutations implicated in ribosomal resistance. The mutations found either lead to amino acid changes in ribosomal protein S12 or alter the primary structure of the 16S rRNA. The 16S rRNA region mutated perturbs a pseudoknot structure in a region which has been linked to ribosomal S12 protein.

Base Sequence↗
PubMed1993

Elimination of contaminating DNA within polymerase chain reaction reagents: implications for a general approach to detection of uncultured pathogens.

Analysis based on comparisons of 16S rRNA sequences provides a rapid and reliable approach to identifying human pathogens. By directing oligonucleotide primers at sequences conserved throughout the eubacterial kingdom, bacterial 16S ribosomal DNA sequences of virtually any member of the eubacterial kingdom can be amplified by polymerase chain reaction and subsequently analyzed by sequence determination. Indeed, automated systems for broad-range amplification, sequencing, and data analysis are now feasible and may form the basis of the next generation of automated microbial identification systems. However, identification of pathogens by this strategy is hampered by the frequent contamination of reagents used for the amplification reaction, in particular Taq polymerase, with exogenous bacterial DNA. Here, we describe detailed investigations on the use of 8-methoxypsoralen and long-wave UV light to eliminate contaminating DNA in polymerase chain reaction reagents. The clinical utility of the developed procedure was demonstrated in a case of paucibacillary osteomyelitis, for which no specific bacterial agent had been cultured.

Bacteria↗
PubMed1992

Multiple nucleic acid labeling and rainbow detection.

A method which allows discrete nucleic acid sequences to be detected with differently colored hybridization signals on the same blot involving only a single hybridization step is described. Nucleic acid probes labeled with digoxigenin, fluorescein, or biotin are hybridized simultaneously to immobilized target nucleic acids. Differential colorimetric detection is carried out in consecutive alkaline phosphatase-based immunoassays with one of three 3-hydroxy-2-naphthoic acid anilide phosphate/diazonium salt combinations as substrate. Each label is visualized by a different color precipitate (green, red, and blue) directly on the membrane. We demonstrate the use of this method in multicolor plasmid mapping, detection of different genomic sequences on a single Southern blot, discrimination of transcription levels in a Northern blot, and colony screening. Advantages and limitations of the method, as well as further applications, are discussed.

Alkaline Phosphatase↗
PubMed1991

Low-dose chymotrypsin treatment inhibits neutrophil migration into sites of inflammation in vivo: effects on Mac-1 and MEL-14 adhesion protein expression and function.

Antibody blocking studies in the mouse suggest that the MEL-14 antigen is involved in neutrophil-endothelial cell interactions and may be important in neutrophil extravasation to sites of inflammation in vivo. We recently showed that chemotactic factor activation causes a rapid (within minutes) shedding of a large fragment of the MEL-14 antigen from the surface of neutrophils. We report here that chymotrypsin, at low doses (0.1 units/1 x 10(6) cells), but not trypsin, elastase, or collagenase, causes an activation-independent rapid loss (greater than 90%) of the MEL-14 antigen from the surface of murine neutrophils. Under the same treatment conditions chymotrypsin has no effect on the expression of four other neutrophil surface antigens, including the Mac-1 adhesion protein. Chymotrypsin treatment has no effect on neutrophil adhesion to plastic, migration to C5a, regulation of the Mac-1 antigen, but causes a greater than 95% reduction in neutrophil binding to high endothelial venules (HEV) in peripheral lymph nodes measured in the ex vivo frozen section HEV binding assay. The level of inhibition of neutrophil adhesion to HEV was comparable to that seen with the MEL-14 antibody. This experimental system allows us for the first time to specifically examine the consequences of removing the MEL-14 antigen from the surface of neutrophils on function in vivo. We show that treatment with chymotrypsin blocks greater than 85% of the ability of neutrophils injected back into the animal to home to the inflamed peritoneum. In similar in vivo experiments the MEL-14 antibody blocks neutrophil homing by 60-70%. These results further support the importance of the MEL-14 antigen in neutrophil extravasation in vivo and indicate that chymotrypsin could be useful in examining the molecular mechanisms involved in extravasation of leukocytes into a variety of diverse tissue sites of inflammation.

Animals↗