TY - JOUR
T1 - Differential enzymatic activity of common haplotypic versions of the human acidic mammalian chitinase protein
AU - Seibold, Max A.
AU - Reese, Tiffany A.
AU - Choudhry, Shweta
AU - Salam, Muhammad T.
AU - Beckman, Kenny
AU - Eng, Celeste
AU - Atakilit, Amha
AU - Meade, Kelley
AU - Lenoir, Michael
AU - Watson, H. Geoffrey
AU - Thyne, Shannon
AU - Kumar, Rajesh
AU - Weiss, Kevin B.
AU - Grammer, Leslie C.
AU - Avila, Pedro
AU - Schleimer, Robert P.
AU - Fahy, John V.
AU - Rodriguez-Santan, Jose
AU - Rodriguez-Cintron, William
AU - Boot, Rolf G.
AU - Sheppard, Dean
AU - Gilliland, Frank D.
AU - Locksley, Richard M.
AU - Burchard, Esteban G.
PY - 2009/7/17
Y1 - 2009/7/17
N2 - Mouse models have shown the importance of acidic mammalian chitinase activity in settings of chitin exposure and allergic inflammation. However, little is known regarding genetic regulation of AMCase enzymatic activity in human allergic diseases. Resequencing the AMCase gene exons we identified 8 non-synonymous single nucleotide polymorphisms including three novel variants (A290G, G296A, G339T) near the gene area coding for the enzyme active site, all in linkage disequilibrium. AMCase protein isoforms, encoded by two gene-wide haplotypes, and differentiated by these three single nucleotide polymorphisms, were recombinantly expressed and purified. Biochemical analysis revealed the isoform encoded by the variant haplotype displayed a distinct pH profile exhibiting greater retention of chitinase activity at acidic and basic pH values. Determination of absolute kinetic activity found the variant isoform encoded by the variant haplotype was 4-, 2.5-, and 10-fold more active than the wild type AMCase isoform at pH 2.2,4.6, and 7.0, respectively. Modeling of the AMCase isoforms revealed positional changes in amino acids critical for both pH specificity and substrate binding. Genetic association analyses of AMCase haplotypes for asthma revealed significant protective associations between the variant haplotype in several asthma cohorts. The structural, kinetic, and genetic data regarding the AMCase isoforms are consistent with the Th2-priming effects of environmental chitin and a role for AMCase in negatively regulating this stimulus.
AB - Mouse models have shown the importance of acidic mammalian chitinase activity in settings of chitin exposure and allergic inflammation. However, little is known regarding genetic regulation of AMCase enzymatic activity in human allergic diseases. Resequencing the AMCase gene exons we identified 8 non-synonymous single nucleotide polymorphisms including three novel variants (A290G, G296A, G339T) near the gene area coding for the enzyme active site, all in linkage disequilibrium. AMCase protein isoforms, encoded by two gene-wide haplotypes, and differentiated by these three single nucleotide polymorphisms, were recombinantly expressed and purified. Biochemical analysis revealed the isoform encoded by the variant haplotype displayed a distinct pH profile exhibiting greater retention of chitinase activity at acidic and basic pH values. Determination of absolute kinetic activity found the variant isoform encoded by the variant haplotype was 4-, 2.5-, and 10-fold more active than the wild type AMCase isoform at pH 2.2,4.6, and 7.0, respectively. Modeling of the AMCase isoforms revealed positional changes in amino acids critical for both pH specificity and substrate binding. Genetic association analyses of AMCase haplotypes for asthma revealed significant protective associations between the variant haplotype in several asthma cohorts. The structural, kinetic, and genetic data regarding the AMCase isoforms are consistent with the Th2-priming effects of environmental chitin and a role for AMCase in negatively regulating this stimulus.
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U2 - 10.1074/jbc.M109.012443
DO - 10.1074/jbc.M109.012443
M3 - Article
C2 - 19435888
AN - SCOPUS:67749096192
SN - 0021-9258
VL - 284
SP - 19650
EP - 19658
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 29
ER -