Abstract
We identified two regions of human LCAT (hLCAT) that when mutated separately to the corresponding rat sequence (E149A and Y292H/W294F) and transiently expressed in COS-1 cells increased phospholipase A2 (PLA2) activity by 5.5- and 2.8-fold, respectively, and increased cholesteryl ester (CE) formation by 2.9- and 1.4-fold, respectively, relative to hLCAT using substrate particles containing 1-16:0,2-20:4-sn-glycero-3-phosphocholine (PAPC). In contrast, both activities with 1-16:0,2-18:1-sn-glycero-3-phosphocholine (POPC) substrate were similar among the three LCAT proteins. The triple mutant (E149A/Y292H/W294F) had increased PLA2 activity with PAPC similar to that observed with the E149A mutation alone; however, unlike E149A, the triple mutant demonstrated a 50% decrease in activity with POPC for both PLA2 activity and CE formation, suggesting an interaction between the two regions of LCAT. Additional mutagenesis studies demonstrated that W294F, but not Y292H, increased PLA2 activity by 3-fold with PAPC without affecting activity with POPC. The E149A/W294F double mutation mimicked the LCAT activity phenotype of the triple mutant (more activity with PAPC, less with POPC). In conclusion, separate mutation of two amino acids in hLC...Continue Reading
References
Sep 1, 1976·The Journal of Clinical Investigation·R L HamiltonR J Havel
Oct 1, 1976·Journal of Biochemistry·U Piran, T Nishida
Jun 1, 1991·Lipids·D Grove, H J Pownall
Feb 22, 1990·Nature·F K WinklerW Hunziker
Apr 15, 1989·Gene·S N HoL R Pease
Apr 3, 1987·Biochimica Et Biophysica Acta·M JauhiainenP J Dolphin
Feb 25, 1972·Biochemical and Biophysical Research Communications·C J FieldingP E Fielding
Jan 18, 1972·Biochemistry·D S Sgoutas
May 19, 1995·The Journal of Biological Chemistry·S E SzedlacsekT Nishida
Nov 15, 1994·Biochemistry·P V SubbaiahF Paltauf
Nov 17, 1994·Biochimica Et Biophysica Acta·P V SubbaiahP H Pritchard
Mar 1, 1994·Lipids·M H Nouri-SorkhabiP W Kuchel
Jan 3, 1997·The Journal of Biological Chemistry·J WangJ S Parks
Dec 12, 1997·European Journal of Biochemistry·F PeelmanM Rosseneu
Apr 16, 1998·Protein Science : a Publication of the Protein Society·F PeelmanM Rosseneu
Jan 26, 1999·Methods in Molecular Biology·J S ParksJ W Furbee
Apr 8, 1999·Progress in Lipid Research·A Jonas
Dec 9, 2000·Biochimica Et Biophysica Acta·A Jonas
Aug 1, 1959·Canadian Journal of Biochemistry and Physiology·E G BLIGH, W J DYER
Jun 1, 1964·Archives of Biochemistry and Biophysics·O W PORTMAN, M SUGANO
Nov 26, 2003·Biochemistry·Yue ZhaoJohn S Parks