Synthesis of a new water-soluble rhodamine derivative and application to protein labeling and intracellular imaging

被引:34
作者
Bandichhor, Rakeshwar
Petrescu, Anca D.
Vespa, Aude
Kier, Ann B.
Schroeder, Friedhelm
Burgess, Kevin
机构
[1] Texas A&M Univ, Dept Chem, College Stn, TX 77842 USA
[2] Texas A&M Univ, TVMC, Dept Physiol, College Stn, TX 77881 USA
[3] Texas A&M Univ, TVMC, Dept Pharmacol, College Stn, TX 77881 USA
[4] Texas A&M Univ, TVMC, Dept Pathobiol, College Stn, TX 77881 USA
关键词
D O I
10.1021/bc0601424
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Synthesis of a new fluorescent rhodamine derivative, dye 1, is reported. This probe is different from other rhodamines insofar as it has several (four) carboxylic acid functionalities to promote water solubility and facilitate conjugation to proteins. It also has an aryl bromide functionality that could, in principle, be used to further functionalize the system for specialized applications. Dye 1 was conjugated to a model protein called ACBP (acyl-CoA binding protein). The properties of this conjugate were tested to establish that the label does not significantly perturb the binding function of the protein to its natural ligand in vitro and to confirm that its secondary structure was not significantly perturbed (circular dichroism). Experiments were performed to test if the labeled protein could be imported into living COS-7 cells (using the Chariot-peptide delivery system) and, if so, to observe, via fluorescence microscopy, which of the labeled protein was able to migrate to the nucleus, as expected for ACBP in cells. In the event, all these postulates were confirmed.
引用
收藏
页码:1219 / 1225
页数:7
相关论文
共 18 条
[1]   CELLULAR AUTOFLUORESCENCE - IS IT DUE TO FLAVINS [J].
BENSON, RC ;
MEYER, RA ;
ZARUBA, ME ;
MCKHANN, GM .
JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY, 1979, 27 (01) :44-48
[2]   Energy transfer cassettes based on BODIPY® dyes [J].
Burghart, A ;
Thoresen, LH ;
Chen, J ;
Burgess, K ;
Bergström, F ;
Johansson, LBÅ .
CHEMICAL COMMUNICATIONS, 2000, (22) :2203-2204
[3]   Membrane charge and curvature determine interaction with acyl-CoA binding protein (ACBP) and fatty acyl-CoA targeting [J].
Chao, H ;
Martin, GG ;
Russell, WK ;
Waghela, SD ;
Russell, DH ;
Schroeder, F ;
Kier, AB .
BIOCHEMISTRY, 2002, 41 (33) :10540-10553
[4]   Acyl coenzyme A binding protein - Conformational sensitivity to long chain fatty acyl-CoA [J].
Frolov, A ;
Schroeder, F .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1998, 273 (18) :11049-11055
[5]   Fluorescent, through-bond energy transfer cassettes for Labeling multiple biological molecules in one experiment [J].
Jiao, GS ;
Thoresen, LH ;
Burgess, K .
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 2003, 125 (48) :14668-14669
[6]   Microwave-assisted syntheses of regioisomerically pure bromorhodamine derivatives [J].
Jiao, GS ;
Castro, JC ;
Thoresen, LH ;
Burgess, K .
ORGANIC LETTERS, 2003, 5 (20) :3675-3677
[7]  
JOSE J, 2006, IN PRESS J ORG CHEM
[8]   CLEAVAGE OF STRUCTURAL PROTEINS DURING ASSEMBLY OF HEAD OF BACTERIOPHAGE-T4 [J].
LAEMMLI, UK .
NATURE, 1970, 227 (5259) :680-+
[9]   MEASUREMENT OF COLOCALIZATION OF OBJECTS IN DUAL-COLOR CONFOCAL IMAGES [J].
MANDERS, EMM ;
VERBEEK, FJ ;
ATEN, JA .
JOURNAL OF MICROSCOPY-OXFORD, 1993, 169 :375-382
[10]  
Mitskyavichius VY, 1996, KHIM GETEROTSIKL+, P1255