title: Non-conservative surface decoration of hemoglobin: influence of neutralization of positive charges at PEGylation sites on molecular and functional properties of PEGylated hemoglobin.
abstract: High hydrodynamic volume, high viscosity and high colloidal osmotic pressure (COP) of PEGylated hemoglobin (Hb) have been suggested to neutralize the vasoactivity of acellular Hb. Consequences of non-conservative PEGylation (positive charge of the amino groups at the PEGylation sites is neutralized) using succinimidyl-ester of propionic acid PEG5K on the properties of PEGylated Hb have now been investigated. Non-conservative PEGylation of Hb leads to a much higher increase in the COP and viscosity of Hb than conservative extension arm facilitated (EAF) PEGylation of Hb. Introduction of alphaalpha-fumaryl crosslinking decreased the COP of non-conservative PEGylated Hb by stabilization of interdimeric interactions. Compared to the EAF-PEGylated alphaalpha-fumaryl Hb, non-conservative PEGylated product shows a comparable COP and higher viscosity. Conservative PEGylation of alphaalpha-fumaryl Hb by reductive alkylation chemistry does not increase the COP to this level, but enhanced the molecular volume and viscosity comparable to EAF-PEGylated product. Thus, the molecular properties of PEGylated Hb can be fine tuned using different PEGylation platforms and provide a unique opportunity for the design of second generation PEGylated Hbs.
authors.affiliation: Department of Physiology and Biophysics, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
authors.forename: Dongxia
authors.initials: D
authors.lastname: Li
authors.forename: Tao
authors.initials: T
authors.lastname: Hu
authors.forename: Belur N
authors.initials: BN
authors.lastname: Manjula
authors.forename: Seetharama A
authors.initials: SA
authors.lastname: Acharya
mesh_terms: D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003102:Colloids; D006441:Hemoglobin A / Q000737:chemistry / Q000627:therapeutic use*; D006454:Hemoglobins / Q000737:chemistry* / Q000627:therapeutic use; D006801:Humans; D010449:Peptide Mapping; D011092:Polyethylene Glycols / Q000737:chemistry* / Q000378:metabolism / Q000627:therapeutic use; D011489:Protein Denaturation; D013499:Surface Properties; D014783:Viscosity
chemical_list: D003102:Colloids; D006454:Hemoglobins; D011092:Polyethylene Glycols; D006441:Hemoglobin A