Synthesis and structural characterization of carboxyethylpyrrole-modified proteins: mediators of age-related macular degeneration.
Protein modifications in which the epsilon-amino group of lysyl residues is incorporated into a 2-(omega-carboxyethyl)pyrrole (CEP) are mediators of age-related macular degeneration (AMD). They promote both angiogenesis into the retina ('wet AMD') and geographic retinal atrophy ('dry AMD'). Blood levels of CEPs are biomarkers for clinical prognosis of the disease. To enable mechanistic studies of their role in promoting AMD, for example, through the activation of B- and T-cells, interaction with receptors, or binding with complement proteins, we developed an efficient synthesis of CEP derivatives, that is especially effective for proteins. The structures of tryptic peptides derived from CEP-modified proteins were also determined. A key finding is that 4,7-dioxoheptanoic acid 9-fluorenylmethyl ester reacts with primary amines to provide 9-fluorenylmethyl esters of CEP-modified proteins that can be deprotected in situ with 1,8-diazabicyclo[5.4.0]undec-7-ene without causing protein denaturation. The introduction of multiple CEP-modifications with a wide variety of CEP:protein ratios is readily achieved using this strategy.
Department of Chemistry, Case Western Reserve University, Cleveland, OH 44106-7078, United States.
Liang
L
Lu
Xiaorong
X
Gu
Li
L
Hong
James
J
Laird
Keeve
K
Jaffe
Jaewoo
J
Choi
John
J
Crabb
Robert G
RG
Salomon
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000378:metabolism; D019921:Biotinylation; D004195:Disease Models, Animal; D008268:Macular Degeneration / Q000175:diagnosis*; D051379:Mice; D010455:Peptides / Q000138:chemical synthesis* / Q000302:isolation & purification / Q000378:metabolism; D011489:Protein Denaturation; D011758:Pyrroles / Q000138:chemical synthesis* / Q000737:chemistry; D012160:Retina / Q000378:metabolism; D012709:Serum Albumin / Q000737:chemistry* / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism
D000906:Antibodies; D010455:Peptides; D011758:Pyrroles; D012709:Serum Albumin; D014357:Trypsin