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Research /Teaching Interests:
Research deals with proteolytic enzymes and their inhibitors, including protein purification, characterization, sequencing, structure function relationships, kinetics, and regulation. Particular emphasis is placed on the roles of proteases and inhibitors in the pathogenesis of human diseases. Work on the structure of human alpha-1-proteinase inhibitor led to the discovery of its active site and we showed that oxidation of a methionine residue in the inhibitory site of α1-PI caused the loss of elastase inhibitory
activity. This finding resulted in the hypothesis that oxidants can lead to a lung-localized inhibitor deficiency. A deficiency of α1-PI, which normally inhibits neutrophil elastase, is known to cause emphysema. This led to studies of the reactions of ozone and nitrogen dioxide on the function of human α1-PI and the secretory leukocyte proteinase inhibitor found in human bronchial mucus, which also inhibits neutrophil elastase. Another long-standing project focuses on the structure and function of human mast cell tryptase, an unusual serine protease that is the most abundant protein in mast cells. Recently, tryptase has been shown to activate protease zymogens, suggesting a critical role in tumor growth and the pathogenesis of arthritis. Finally, tryptase has an anticoagulant function due to the limited proteolysis of blood coagulation proteins, including fibrinogen and high molecular weight kininogen. Currently, research focuses on the expression of recombinant human mast cell and neutrophil serine proteases, as well as human enterokinase and C-reactive protein, using Pichia pastoris as the host organism.
Research Support:
2001-2004 “Recombinant Human Mast Cell Tryptases”, NIAID, NIH R15 AI45549, $126,954.
2007-2011 “Human Cathepsin G: Expression, C-Terminal Processing and Dual Specificity” National Heart Lung and Blood Institute
1R15HL091770 $210,230.
Lab Members:
Eliot Smith – Ph.D. Graduate student
Evan Perry – Undergraduate Honors student
Dustin Wood – Undergraduate – Biological Sciences
Megan Sears – Undergraduate – Biological Sciences
Selected References:
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1:
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Johnson DA.
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Related Articles
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Human mast cell proteases: activity assays using thiobenzyl ester substrates.
Methods Mol Biol. 2006;315:193-202.
PMID: 16110159 [PubMed - indexed for MEDLINE]
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4:
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Kilic F, Johnson DA, Sinensky M.
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Related Articles
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Subcellular localization and partial purification of prelamin A endoprotease: an enzyme which catalyzes the conversion of farnesylated prelamin A to mature lamin A.
FEBS Lett. 1999 Apr 30;450(1-2):61-5.
PMID: 10350058 [PubMed - indexed for MEDLINE]
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6:
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Addington AK, Johnson DA.
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Related Articles
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Inactivation of human lung tryptase: evidence for a re-activatable tetrameric intermediate and active monomers.
Biochemistry. 1996 Oct 22;35(42):13511-8.
PMID: 8885830 [PubMed - indexed for MEDLINE]
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7:
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Little SS, Johnson DA.
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Related Articles
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Human mast cell tryptase isoforms: separation and examination of substrate-specificity differences.
Biochem J. 1995 Apr 15;307 ( Pt 2):341-6.
PMID: 7733867 [PubMed - indexed for MEDLINE]
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11:
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Johnson DA.
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Related Articles
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Effects of ozone and nitrogen dioxide on human lung proteinase inhibitors.
Res Rep Health Eff Inst. 1987;(11):5-25.
PMID: 3268287 [PubMed - indexed for MEDLINE]
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Book Chapters
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