The influence of deletion mutations on phospholipase C-gamma 1 activity.
Phospholipase C-gamma1, a substrate for many growth factor receptor and nonreceptor tyrosine kinases, produces second messenger molecules that are elements of signal transduction pathways related to cell proliferation. The influence of deletion mutations, which do not intrude on the domains required for catalytic function, on the basal activity of this enzyme is reported. Removal of the first 74 amino-terminal residues increases phospholipase C activity, while deletion of the carboxy-terminal 81 residues decreases enzyme activity. Deletion of the SH2-SH2-SH3 central region, which separates the two domains (X, Y) responsible for catalytic function, also increases enzymatic activity. Interestingly, addition of a recombinant SH2-SH2-SH3 fragment of phospholipase C-gamma1 to the holoenzyme inhibits its phospholipase activity at pH 7.0, but not at pH 5.0. However, addition of individual SH2 or SH3 domains does not influence activity of the holoenzyme. All three deletion mutants, in contrast to the holoenzyme, are relatively resistant to V8 proteolysis and activation induced by the epidermal growth factor receptor tyrosine kinase, which require, respectively, specific proteolysis and phosphorylation sites within the SH region. This suggests a conformational change is induced in the SH region by deletion at either the amino- or carboxy-terminus.
Horstman
D A
DA
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee, 37232, USA.
Chattopadhyay
A
A
Carpenter
G
G
D000818:Animals; D002460:Cell Line; D004789:Enzyme Activation / Q000235:genetics; D007313:Insecta; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D051966:Phospholipase C gamma; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D051381:Rats; D017384:Sequence Deletion; D010738:Type C Phospholipases / Q000235:genetics* / Q000378:metabolism*; D018909:src Homology Domains
D007527:Isoenzymes; D010738:Type C Phospholipases; D051966:Phospholipase C gamma