Differential enzymatic activity of common haplotypic versions of the human acidic Mammalian chitinase protein.
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.
Seibold
Max A
MA
Department of Medicine, Howard Hughes Medical Institute, University of California, San Francisco, California 94143, USA. seiboldm@njc.org
Reese
Tiffany A
TA
Choudhry
Shweta
S
Salam
Muhammad T
MT
Beckman
Kenny
K
Eng
Celeste
C
Atakilit
Amha
A
Meade
Kelley
K
Lenoir
Michael
M
Watson
H Geoffrey
HG
Thyne
Shannon
S
Kumar
Rajesh
R
Weiss
Kevin B
KB
Grammer
Leslie C
LC
Avila
Pedro
P
Schleimer
Robert P
RP
Fahy
John V
JV
Rodriguez-Santana
Jose
J
Rodriguez-Cintron
William
W
Boot
Rolf G
RG
Sheppard
Dean
D
Gilliland
Frank D
FD
Locksley
Richard M
RM
Burchard
Esteban G
EG
D001741:Black or African American / Q000235:genetics; D000818:Animals; D001249:Asthma / Q000208:ethnology / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D002384:Catalysis; D002460:Cell Line; D002688:Chitinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004187:Disaccharides / Q000737:chemistry / Q000378:metabolism; D005787:Gene Frequency; D005838:Genotype; D006239:Haplotypes; D006630:Hispanic or Latino / Q000235:genetics; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007313:Insecta; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D015810:Linkage Disequilibrium; D008800:Mexico; D015394:Molecular Structure; D020641:Polymorphism, Single Nucleotide; D017434:Protein Structure, Tertiary; D011647:Puerto Rico; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
D004187:Disaccharides; D007527:Isoenzymes; D011994:Recombinant Proteins; C032438:chitobiose; C499254:CHIA protein, human; D002688:Chitinases