Consequences of seven novel mutations on the expression and structure of keratinocyte transglutaminase.

We report the molecular characterization of seven new keratinocyte transglutaminase mutations (R315C, S358R, V379L, G473S, R687C, deletion Delta679-696, R127Stop) found in lamellar ichthyosis patients. Arg-315, Ser-358, Val-379, and Gly-473 are highly conserved residues in transglutaminases while Arg-687 and Delta679-696 are not. All mutations strongly decreased transglutaminase activity and protein levels. The mutation R127Stop diminished the amount of mRNA. Structural analysis of these mutations based on the factor XIII A-subunit crystal structure demonstrated that Arg-315, Ser-358, Val-379, and Gly-473 are located in the catalytic core domain, and Arg-687 and the deletion are in the beta-barrel domains. The side chains of amino acids Arg-315, Ser-358, and Gly-473 make ionic and hydrogen bonds important for folding and structural stability of the enzyme but are not directly involved in catalysis. Val-379 is two amino acids away from the active site cysteine, and its change into leucine disturbs the active site structure. The decreased activity and protein level after expression of the R687C and Delta679-696 TGK cDNA in TGK negative keratinocytes excluded that they are polymorphisms. These results identify important amino acids in the central core domain of transglutaminases and show that the C-terminal end influences the structural and functional integrity of TGK.