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post-doc/transpositional recombination Date sent:

Harri Savilahti harri.savilahti at helsinki.fi
Tue Jan 28 04:49:27 EST 1997


UNIVERSITY OF HELSINKI
post doctoral position for 3 years 

TRANSPOSITIONAL DNA RECOMBINATION   


Send an application with a CV and two references
Harri Savilahti, Ph.D.
Institute of Biotechnology 
Viikinkaari 9
00014 University of Helsinki
FINLAND 
tel. int-358-9-708 59516, FAX int-358-9-708 59366
E-mail: harri.savilahti at helsinki.fi




GENERAL BACKGROUND

During the last few decades it has become clear that genomes are not
rigid and stable. Segments of DNA are moved, inverted, duplicated and
extensively multiplied within living cells; sometimes seemingly
purposefully (e. g. switching of immunoglobulin genes), sometimes
fortuitously (e. g. many of the transposition reactions). Very often
the results are harmful to the organism (e.g. many of the
rearrangements that ultimately lead to cancer). Many viruses modify
genomes by integrating their DNA into the genome of their host
organisms; the consequences can be disastrous (e. g. AIDS caused by
HIV). DNA recombination is a term that describes all rearragements
within the genome. However, the usage of this term was originally
limited to genetic recombination during cell division (homologous
recombination). The mechanim(s) of recombination are essential for
living cells and as such an important area of study. In addition,
understanding of different kind of recombination reactions apparently
creates possibilities for applications, especially in medicine but
also in many fields of biotechnology/genetic engineering. Future gene
therapy and plant breeding are dependent on mastering such reactions. 


DNA transposition

DNA transposition is a recombination reaction in which a DNA element
(a transposon) moves from one location to another in the genome of its
host organism. The underlying molecular mechanisms are universal from
lower prokaryotes to higher eukaryotes. At the molecular level
retrovirus integration is also a transposition reaction. In this
reaction specific protein molecules utilizing tightly controlled
biochemical reactions excise a piece of DNA and splice it to another
location in the genome. 

Bacteriophage Mu multiplies by using efficient transposition;
transposition is facilitated by transposition complexes

Bacteriophage Mu is a virus that replicates by using the mechanisms of
DNA transposition. Of the known transposition systems it is by far the
most efficient and its transposition reaction was the first to be
established in vitro  Mu transposition reactions proceed in the
context of specific protein-DNA complexes, in the heart of which is
the critical functional and structural component, the transposase. In
Mu this protein is MuA and it catalyzes the DNA cleavage and joining
reactions. It functions in transposition as a tetramer which also
synapses the two transposon ends.

We have shown that under certain reaction conditions fully functional
Mu transposition complexes (transpososomes) can be formed in vitro by
using short synthetic Mu-specific DNA segments and MuA protein. When
the active transposition complex has been formed it can react, in
principle, with any other DNA molecule (target DNA). Because we now
can make transposition complexes from its minimal components in vitro 
this offers an excellent opportunity to study the governing principles
in transposition at the molecular level.

Recent publications:

Baker, T. A., Mizuuchi, M., Savilahti, H. and Mizuuchi, K. 			1993:
Division of labor among monomers within the 			Mu transposase
tetramer. -Cell 74: 723-733.

Clubb, R. T., Omichinski, J. G., Savilahti, H., Mizuuchi, 			K.,
Gronenborn, A. M. and Clore, G. M. 1994: A 			novel class of winged
helix-turn-helix protein: 			The DNA binding domain of Mu transposase.
-Structure 2: 1041-1048.

Savilahti, H., Rice, P. A. and Mizuuchi, K. 1995: The 			phage Mu
transpososome core: DNA requirements for 			assembly and function.
-EMBO Journal 19: 4893-4903. 

Clubb, R. T., Mizuuchi, M., Huth, J. R., Omichinski, J. G.,
			Savilahti, H., Mizuuchi, K., Clore, G. M. and 			Gronenborn, A. M.
1996: The wing of the enhancer 			binding domain of Mu phage
transposase is flexible 			and essential for efficient transposition.
-Proc. Natl. Acad. Sci. USA 93:1146-1150. 

Savilahti, H. and Mizuuchi, K. 1996: Mu transpositional
			recombination: donor DNA cleavage and strand 			transfer in trans
by the MuA transposase. -Cell 85:271-280.






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