A core–shell
ensemble of bovine hemoglobin (Hb) and human serum albumin (HSA) is
an artificial O2 carrier as a red blood cell substitute.
This protein particle is created by covalent wrapping of a carbonyl
Hb with HSAs: HbR–HSA
3
cluster, where HbR signifies the use of carbonyl Hb (relaxed
(R) state conformation) as a starting material. The HbR–HSA
3
cluster exhibits high O2 affinity and low cooperativity. Analysis of the quaternary
structure of the central HbR in the cluster revealed that
its high O2 affinity is attributed to the physically immobile
HbR nucleus. Circular dichroism and UV–vis absorption
spectroscopy showed that the structure of deoxy HbR core
closely resembles the R-state. The crystal structure of Lys-modified
carbonyl HbR was superimposed on that of carbonyl Hb. These
results imply that chemical modifications of the surface Lys groups
and Cys-93(β) of the carbonyl Hb with cross-linking agent interfered
in the quaternary structure movement from the R-state to tense (T)
state. As expected, coupling of deoxy Hb (T-state) with HSAs yielded
HbT–HSA
3
cluster having
low O2 affinity. The mixing of HbR–HSA
3
and HbT–HSA
3
clusters conferred a tailor-made formulation of
artificial O2 carrier with a desired O2 affinity
(P
50).
Myoglobin combined with human serum albumin (Mb-HSA) can be produced using yeast Pichia pastoris as a host strain, with secretion into the culture medium. This Mb-HSA fusion protein possesses identical O 2 binding affinity to that of naked Mb. The Mb unit is reconstituted with a zinc(II) protoporphyrin IX, yielding (zinc substituted Mb)-HSA, ZnMb-HSA. The photophysical property and singlet O 2 generation ability of ZnMb-HSA are equivalent to those of ZnMb. In vitro cell experiments revealed that ZnMb-HSA acts as a superior photosensitizer for photodynamic cancer therapy. It is noteworthy that ZnMb-HSA shows long circulation lifetime in vivo.
A hemoglobin (Hb) wrapped covalently by three human serum albumins (HSAs) is a triangular protein cluster designed as an artificial O-carrier and red blood cell substitute. We report the structural insights into this Hb-HSA cluster in aqueous medium revealed by 3D reconstruction based on cryogenic transmission electron microscopy (cryo-TEM) data and small-angle X-ray scattering (SAXS) measurements. Cryo-TEM observations showed individual particles with approximately 15 nm diameter in the vitrified ice layer. Subsequent image processing and 3D reconstruction proved the expected spatial arrangements of an Hb in the center and three HSAs at the periphery. SAXS measurements demonstrated the monodispersity of the Hb-HSA cluster having a molecular mass of 270 kDa. The pair-distance distribution function suggested the existence of oblate-like particles with a maximum dimeter of ∼17 nm. The supramolecular 3D structure reconstructed from the SAXS intensity using an ab initio procedure was similar to that obtained from cryo-TEM data.
We describe the synthesis, photophysical properties, and photodynamic activity of a methemoglobin (metHb) wrapped covalently by human serum albumins (HSAs) incorporating protoporphyrin IX (PPIX): a metHb‐HSA3‐PPIX2 cluster. The metHb core catalyzes H2O2 disproportionation to generate O2 in tumor tissue. The HSA3‐PPIX2 shell acts as a photosensitizer for 1O2 formation. The metHb‐HSA3‐PPIX2 cluster acts as a dual functional protein drug for photodynamic therapy.
Photodynamic therapy (PDT) is a non-invasive cancer treatment using reactive oxygen species (ROS) generated by light irradiation. Excited state photosensitizer produces active radicals (eg. O2•– and OH•) by electron transfer...
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