J. Biol. Chem., Vol. 266, Issue 17, 10781-10786, 06, 1991
Structural and catalytic properties of a deletion derivative (delta 133- 157) of Escherichia coli adenylate kinase
T Rose, M Brune, A Wittinghofer, K Le Blay, WK Surewicz, HH Mantsch, O Barzu and AM Gilles
Unite de Biochimie des Regulations Cellulaires, Institut Pasteur, Paris, France.
Escherichia coli adenylate kinase (AKe) as well as the enzyme from yeast
and mitochondria differs from the muscle cytosolic variant (AK1) by an
insertion of 25 amino acid residues that are missing in AK1. The extra
sequence, highly homologous in "large" size variants, is situated between
residues 133 and 157 in AKe. Removal of 25 codons in the corresponding adk
gene resulted in expression of a modified form of adenylate kinase (delta
133-157 AKe) which still conserved 7% of the maximal activity of the
wild-type protein. The apparent Km for nucleotide substrates was increased
by a factor of 4.6 (ADP), 23 (ATP) or 43 (AMP) in delta 133-157 AKe when
compared with the wild-type enzyme. The secondary structure of delta
133-157 AKe, as well as its thermal stability were very similar to the
parent protein. However, the deleted protein was much more sensitive than
the wild-type enzyme to inactivation by trypsin. Sodium dodecyl
sulfate-polyacrylamide gel electrophoresis analysis of trypsin digested
delta 133-157 AKe revealed accumulation of several well defined fragments
which were not observed in the case of wild-type enzyme. We conclude that
the additional sequence, although necessary for expression of full activity
in AKe, is not critical for catalysis. It is perhaps responsible for
interaction of enzyme with other cellular components although a different
mechanism of water shielding for large and small size variants of AK can be
also envisaged.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
