INVESTIGADORES
GUBERMAN Alejandra Sonia
artículos
Título:
Screening for and characterization of phospholipase A1 hypersecretory mutants of Tetrahymena thermophila. Appl. Microbiol. Biotechnol
Autor/es:
HARTMANN, M., GUBERMAN, A., FLORIN-CHRISTENSEN, M. TIEDTKE, A.
Revista:
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
Referencias:
Año: 2000 vol. 54 p. 390 - 396
ISSN:
0175-7598
Resumen:
We have described a procedure for the isolation of mutants of
Tetrahymena thermophila with hypersecretion of phospholipase A1 (PLA1).
Using random chemical mutagenesis, uniparental cytogamy, genetic
crossing and a new, fast and effective screening procedure, four
PLA1-hypersecretory mutants were isolated. The screening procedure is
based on the formation of a halo appearing around cylindrical holes in
a lecithin-containing agar plate filled with cell-free supernatants.
About 3,940 clones were tested with this procedure in primary screening
for hypersecretory features, of which 60 putative hypersecretory
mutants were isolated, subcloned and tested in a secondary screening.
Of these, four selected mutants showed 1.8-2.2 more PLA1 activity in
the cell-free supernatants compared to the wild-type strain CU 438.1.
Hypersecretion was only observable for PLA1; no increased activity for
two other lysosomal enzymes could be detected. These hypersecretory
mutants of T. thermophila can be very useful for increasing the yield
of PLA1 in fermentation processes. This is particularly relevant
because, in contrast to other phospholipases, PLA1 is not available on
the commercial market for fine chemicals and little is known about the
role of PLA1 in cell signaling and metabolism.