title: Altered specificity of Hint-W123Q supports a role for Hint inhibition by ASW in avian sex determination.
abstract: Hint is a universally conserved, dimeric AMP-lysine hydrolase encoded on the avian Z chromosome. Tandemly repeated on the female-specific W chromosome, Asw encodes a potentially sex-determining, dominant-negative Hint dimerization partner whose substrate-interacting residues were specifically altered in evolution. To test the hypothesis that Gln127 of Asw is responsible for depression and/or alteration of Hint enzyme activity, a corresponding mutant was created in the chicken Hint homodimer, and a novel substrate was developed that links reversal of AMP-lysine modification to aminomethylcoumarin release. Strikingly, the Hint-W123Q substitution reduced k(cat)/K(m) for AMP-lysine hydrolysis 17-fold, while it increased specificity for AMP-para-nitroaniline hydrolysis by 160-fold. The resulting 2,700-fold switch in enzyme specificity suggests that Gln127 could be the dominant component of Asw dominant negativity in avian feminization.
authors.affiliation: Department of Genetics, Norris Cotton Cancer Center, Dartmouth Medical School, Lebanon, New Hampshire 03756, USA.
authors.forename: Kristen P
authors.initials: KP
authors.lastname: Parks
authors.forename: Heather
authors.initials: H
authors.lastname: Seidle
authors.forename: Nathan
authors.initials: N
authors.lastname: Wright
authors.forename: Jeffrey B
authors.initials: JB
authors.lastname: Sperry
authors.forename: Pawel
authors.initials: P
authors.lastname: Bieganowski
authors.forename: Konrad
authors.initials: K
authors.lastname: Howitz
authors.forename: Dennis L
authors.initials: DL
authors.lastname: Wright
authors.forename: Charles
authors.initials: C
authors.lastname: Brenner
mesh_terms: D000249:Adenosine Monophosphate / Q000737:chemistry; D000814:Aniline Compounds / Q000737:chemistry; D000818:Animals; D030161:Avian Proteins; D001717:Birds; D002352:Carrier Proteins / Q000502:physiology*; D002645:Chickens; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D019143:Evolution, Molecular; D020224:Expressed Sequence Tags; D005260:Female; D005799:Genes, Dominant; D005973:Glutamine / Q000737:chemistry; D006867:Hydrolases / Q000737:chemistry* / Q000235:genetics*; D006868:Hydrolysis; D007621:Karyotyping; D007700:Kinetics; D008239:Lysine / Q000737:chemistry; D008297:Male; D008956:Models, Chemical; D009154:Mutation; D017434:Protein Structure, Tertiary; D012730:Sex Chromosomes / Q000648:ultrastructure*; D019849:Sex Determination Processes; D013379:Substrate Specificity
chemical_list: D000814:Aniline Compounds; D030161:Avian Proteins; D002352:Carrier Proteins; D005973:Glutamine; C019498:4-nitroaniline; D000249:Adenosine Monophosphate; D006867:Hydrolases; C510383:AMP-lysine hydrolase, chicken; D008239:Lysine