title: Structural and Functional Analysis of Human HtrA3 Protease and Its Subdomains.
abstract: Human HtrA3 protease, which induces mitochondria-mediated apoptosis, can be a tumor suppressor and a potential therapeutic target in the treatment of cancer. However, there is little information about its structure and biochemical properties. HtrA3 is composed of an N-terminal domain not required for proteolytic activity, a central serine protease domain and a C-terminal PDZ domain. HtrA3S, its short natural isoform, lacks the PDZ domain which is substituted by a stretch of 7 C-terminal amino acid residues, unique for this isoform. This paper presents the crystal structure of the HtrA3 protease domain together with the PDZ domain (ΔN-HtrA3), showing that the protein forms a trimer whose protease domains are similar to those of human HtrA1 and HtrA2. The ΔN-HtrA3 PDZ domains are placed in a position intermediate between that in the flat saucer-like HtrA1 SAXS structure and the compact pyramidal HtrA2 X-ray structure. The PDZ domain interacts closely with the LB loop of the protease domain in a way not found in other human HtrAs. ΔN-HtrA3 with the PDZ removed (ΔN-HtrA3-ΔPDZ) and an N-terminally truncated HtrA3S (ΔN-HtrA3S) were fully active at a wide range of temperatures and their substrate affinity was not impaired. This indicates that the PDZ domain is dispensable for HtrA3 activity. As determined by size exclusion chromatography, ΔN-HtrA3 formed stable trimers while both ΔN-HtrA3-ΔPDZ and ΔN-HtrA3S were monomeric. This suggests that the presence of the PDZ domain, unlike in HtrA1 and HtrA2, influences HtrA3 trimer formation. The unique C-terminal sequence of ΔN-HtrA3S appeared to have little effect on activity and oligomerization. Additionally, we examined the cleavage specificity of ΔN-HtrA3. Results reported in this paper provide new insights into the structure and function of ΔN-HtrA3, which seems to have a unique combination of features among human HtrA proteases.
authors.affiliation: Department of Biochemistry, Faculty of Biology, University of Gdansk, 80-308 Gdansk, Poland.
authors.forename: Przemyslaw
authors.initials: P
authors.lastname: Glaza
authors.affiliation: Midwest Center for Structural Genomics, Argonne National Laboratory, Argonne, Illinois, IL 60439, United States of America; Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois, IL 60439, United States of America.
authors.forename: Jerzy
authors.initials: J
authors.lastname: Osipiuk
authors.affiliation: Department of Biochemistry, Faculty of Biology, University of Gdansk, 80-308 Gdansk, Poland.
authors.forename: Tomasz
authors.initials: T
authors.lastname: Wenta
authors.affiliation: Department of Biochemistry, Faculty of Biology, University of Gdansk, 80-308 Gdansk, Poland.
authors.forename: Dorota
authors.initials: D
authors.lastname: Zurawa-Janicka
authors.affiliation: Department of Biochemistry, Faculty of Biology, University of Gdansk, 80-308 Gdansk, Poland.
authors.forename: Miroslaw
authors.initials: M
authors.lastname: Jarzab
authors.affiliation: Department of Biochemistry, Faculty of Chemistry, University of Gdansk, 80-308 Gdansk, Poland.
authors.forename: Adam
authors.initials: A
authors.lastname: Lesner
authors.affiliation: Department of Molecular and Cellular Biology, Intercollegiate Faculty of Biotechnology of the University of Gdansk and the Medical University of Gdansk, 80-822 Gdansk, Poland.
authors.forename: Bogdan
authors.initials: B
authors.lastname: Banecki
authors.affiliation: Department of Biochemistry, Faculty of Biology, University of Gdansk, 80-308 Gdansk, Poland.
authors.forename: Joanna
authors.initials: J
authors.lastname: Skorko-Glonek
authors.affiliation: Midwest Center for Structural Genomics, Argonne National Laboratory, Argonne, Illinois, IL 60439, United States of America; Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois, IL 60439, United States of America.
authors.forename: Andrzej
authors.initials: A
authors.lastname: Joachimiak
authors.affiliation: Department of Biochemistry, Faculty of Biology, University of Gdansk, 80-308 Gdansk, Poland.
authors.forename: Barbara
authors.initials: B
authors.lastname: Lipinska
mesh_terms: D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D054731:PDZ Domains / Q000235:genetics / Q000502:physiology*; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry / Q000502:physiology; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D013329:Structure-Activity Relationship
chemical_list: D020033:Protein Isoforms; D021122:Protein Subunits; C474013:HTRA3 protein, human; D012697:Serine Endopeptidases