Lysine 1423 of neurofibromin (neurofibromatosis type I gene product [NF1]) plays 
a crucial role in the function of NF1. Mutations of this lysine were detected in 
samples from a neurofibromatosis patient as well as from cancer patients. To 
further understand the significance of this residue, we have mutated it to all 
possible amino acids. Functional assays using yeast ira complementation have 
revealed that lysine is the only amino acid that produced functional NF1. 
Quantitative analyses of different mutant proteins have suggested that their 
GTPase-activating protein (GAP) activity is drastically reduced as a result of a 
decrease in their Ras affinity. Such a requirement for a specific residue is not 
observed in the case of other conserved residues within the GAP-related domain. 
We also report that another residue, phenylalanine 1434, plays an important role 
in NF1 function. This was first indicated by the finding that defective NF1s due 
to an alteration of lysine 1423 to other amino acids can be rescued by a second 
site intragenic mutation at residue 1434. The mutation partially restored GAP 
activity in the lysine mutant. When the mutation phenylalanine 1434 to serine 
was introduced into a wild-type NF1 protein, the resulting protein acquired the 
ability to suppress activated phenotypes of RAS2Val-19 cells. This suppression, 
however, does not involve Ras interaction, since the phenylalanine mutant does 
not stimulate the intrinsic GTPase activity of RAS2Val-19 protein and does not 
have an increased affinity for Ras proteins.