163 211

Cited 5 times in

Novel point mutations attenuate autotaxin activity

Authors
 Eunjin Koh  ;  Russell W Bandle  ;  David D Roberts  ;  Mary L Stracke  ;  Timothy Clair 
Citation
 LIPIDS IN HEALTH AND DISEASE, Vol.8 : 4, 2009 
Journal Title
 LIPIDS IN HEALTH AND DISEASE 
Issue Date
2009
MeSH
Amino Acid Substitution/drug effects ; Cell Movement/drug effects ; Fatty Acids/metabolism ; Humans ; Hydrolysis/drug effects ; Immunoblotting ; Kinetics ; Lysophospholipids/pharmacology ; Multienzyme Complexes/genetics* ; Multienzyme Complexes/metabolism* ; Mutant Proteins/genetics ; Mutant Proteins/metabolism* ; Phosphodiesterase I/genetics* ; Phosphodiesterase I/metabolism* ; Phosphoric Diester Hydrolases ; Point Mutation/genetics* ; Pyrophosphatases/genetics* ; Pyrophosphatases/metabolism* ; Substrate Specificity/drug effects
Keywords
Histidine Residue ; Artificial Substrate ; Migration Response ; Choline Oxidase ; A2058 Human Melanoma Cell
Abstract
BACKGROUND: The secreted enzyme autotaxin (ATX) stimulates tumor cell migration, tumorigenesis, angiogenesis, and metastasis. ATX hydrolyzes nucleotides, but its hydrolysis of lysophospholipids to produce lysophosphatidic acid (LPA) accounts for its biological activities. ATX has been identified only as a constitutively active enzyme, and regulation of its activity is largely unexplored. In spite of its presence in plasma along with abundant putative substrate LPC, the product LPA is found in plasma at unexpectedly low concentrations. It is plausible that the LPA-producing activity of ATX is regulated by its expression and by access to substrate(s). For this reason studying the interaction of enzyme with substrate is paramount to understanding the regulation of LPA production. RESULTS: In this study we determine ATX hydrolytic activities toward several artificial and natural substrates. Two novel point mutations near the enzyme active site (H226Q and H434Q) confer attenuated activity toward all substrates tested. The Vmax for LPC compounds depends upon chain length and saturation; but this order does not differ among wild type and mutants. However the mutant forms show disproportionately low activity toward two artificial substrates, pNpTMP and FS-3. The mutant forms did not significantly stimulate migration responses at concentrations that produced a maximum response for WT-ATX, but this defect could be rescued by inclusion of exogenous LPC. CONCLUSION: H226Q-ATX and H434Q-ATX are the first point mutations of ATX/NPP2 demonstrated to differentially impair substrate hydrolysis, with hydrolysis of artificial substrates being disproportionately lower than that of LPC. This implies that H226 and H434 are important for substrate interaction. Assays that rely on hydrolyses of artificial substrates (FS-3 and pNpTMP), or that rely on hydrolysis of cell-derived substrate, might fail to detect certain mutated forms of ATX that are nonetheless capable of producing LPA in the presence of sufficient exogenous substrate. H420Q-ATX could not be differentiated from WT-ATX, indicating that histidine at position 420 is not required for any of the activities of ATX tested in this study
Files in This Item:
T200906171.pdf Download
DOI
10.1186/1476-511X-8-4
Appears in Collections:
1. College of Medicine (의과대학) > Dept. of Biochemistry and Molecular Biology (생화학-분자생물학교실) > 1. Journal Papers
Yonsei Authors
Koh, Eun Jin(고은진) ORCID logo https://orcid.org/0000-0001-8967-6266
URI
https://ir.ymlib.yonsei.ac.kr/handle/22282913/106060
사서에게 알리기
  feedback

qrcode

Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.

Browse