Presentation Title

Oncogenic Lysyl Oxidase Mutations: Evaluating the Effects of Y203A

Faculty Mentor

Karlo Lopez

Start Date

23-11-2019 8:00 AM

End Date

23-11-2019 8:45 AM

Location

105

Session

poster 1

Type of Presentation

Poster

Subject Area

biological_agricultural_sciences

Abstract

Lysyl oxidase (LOX) is copper-dependent amine oxidase that plays a key role in the crosslinking of collagen and elastin. The enzyme is synthesized as a 50 kDa proenzyme inside the cell and it is exported into the extracellular matrix where it is cleaved by procollagen-C-proteinase into a propeptide and the 29 kDa mature form of the enzyme. Structurally, the enzyme is stable, presumably because it contains five disulfide crosslinkages and a crosslinked cofactor, lysyl tyrosyl quinone (LTQ). In the extracellular matrix, the LOX propeptide acts as a repressor of the ras oncogene while the mature form of the enzyme acts to crosslink collagen and elastin. LOX overexpression increases when a malignant transformation occurs in the cell, leading to increase crosslinking and eventually metastasis. The role that lysyl oxidase plays in cancer is still not well understood. Low solubility and a lack of an overexpression system that produces high yields of active enzyme have hampered the characterization of this enzyme for many years. Recent advances in our laboratory, however, have allowed us to overcome these problems and begin to characterize this enzyme. In particular, we have shown that the solubility and overexpression problem can be overcome and that the constructs generated can be used to determine the roles of specific amino acids. By using a NUS-A solubility tag on the N-terminus, the need for urea has been eliminated without sacrificing activity (0.11 U/mg) or overexpression yield (10 mg of protein per liter of media) in the wildtype. These numbers will be used as baseline in conjunction with percent copper incorporation (68%) and LTQ formation to determine the effects of any mutations. Having developed the methodology to probe the effects of specific amino acids in soluble and active enzyme, this project aims to evaluate the effects of seven mutations commonly found in oncogenic LOX. The mutations were selected from the Catalog of Somatic Mutations in Cancer (COSMIC) which curates data from papers in the scientific literature and large scale experimental screens from the Cancer Genome Project at the Sanger Institute in the United Kingdom. This work focuses on the Y203A mutation.

This document is currently not available here.

Share

COinS
 
Nov 23rd, 8:00 AM Nov 23rd, 8:45 AM

Oncogenic Lysyl Oxidase Mutations: Evaluating the Effects of Y203A

105

Lysyl oxidase (LOX) is copper-dependent amine oxidase that plays a key role in the crosslinking of collagen and elastin. The enzyme is synthesized as a 50 kDa proenzyme inside the cell and it is exported into the extracellular matrix where it is cleaved by procollagen-C-proteinase into a propeptide and the 29 kDa mature form of the enzyme. Structurally, the enzyme is stable, presumably because it contains five disulfide crosslinkages and a crosslinked cofactor, lysyl tyrosyl quinone (LTQ). In the extracellular matrix, the LOX propeptide acts as a repressor of the ras oncogene while the mature form of the enzyme acts to crosslink collagen and elastin. LOX overexpression increases when a malignant transformation occurs in the cell, leading to increase crosslinking and eventually metastasis. The role that lysyl oxidase plays in cancer is still not well understood. Low solubility and a lack of an overexpression system that produces high yields of active enzyme have hampered the characterization of this enzyme for many years. Recent advances in our laboratory, however, have allowed us to overcome these problems and begin to characterize this enzyme. In particular, we have shown that the solubility and overexpression problem can be overcome and that the constructs generated can be used to determine the roles of specific amino acids. By using a NUS-A solubility tag on the N-terminus, the need for urea has been eliminated without sacrificing activity (0.11 U/mg) or overexpression yield (10 mg of protein per liter of media) in the wildtype. These numbers will be used as baseline in conjunction with percent copper incorporation (68%) and LTQ formation to determine the effects of any mutations. Having developed the methodology to probe the effects of specific amino acids in soluble and active enzyme, this project aims to evaluate the effects of seven mutations commonly found in oncogenic LOX. The mutations were selected from the Catalog of Somatic Mutations in Cancer (COSMIC) which curates data from papers in the scientific literature and large scale experimental screens from the Cancer Genome Project at the Sanger Institute in the United Kingdom. This work focuses on the Y203A mutation.