Functional characterization of heme proteins encapsulated in wet nanoporous silica gels.
Myoglobin and hemoglobin were encapsulated in wet, nanoporous silica gels. A rigorous evaluation of the effect of the encapsulation on protein dynamics and function was carried out by measuring the kinetics of carbon monoxide rebinding after nanosecond laser flash-photolysis with transient absorption detection, and the oxygen affinity with absorption microspectrophotometry. The time course of carbon monoxide binding to myoglobin evidenced a strongly enhanced geminate recombination and a faster bimolecular rebinding with respect to solution, whereas T and R quaternary states of hemoglobin exhibited a geminate phase and a bimolecular binding rate very similar to those observed in solution. Oxygen affinity of T-state hemoglobin was found to be close to that observed for the binding of the first oxygen to T-state hemoglobin in solution. Results indicate that some conformational transitions are kinetically restricted, allowing to isolate distinct tertiary and quaternary states. This opens the way to their detailed functional characterization and application to biodevices.
Abbruzzetti
S
S
National Institute for the Physics of Matter (INFM), Department of Physics, University of Parma, Via Parco delle Scienze, 43100 Parma, Italy.
Viappiani
C
C
Bruno
S
S
Bettati
S
S
Bonaccio
M
M
Mozzarelli
A
A
D000042:Absorption; D000818:Animals; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis / Q000737:chemistry; D004339:Drug Compounding / Q000379:methods*; D006420:Hemeproteins / Q000737:chemistry / Q000302:isolation & purification; D006454:Hemoglobins / Q000737:chemistry* / Q000302:isolation & purification; D006801:Humans; D008567:Membranes, Artificial; D008968:Molecular Conformation; D009211:Myoglobin / Q000737:chemistry* / Q000302:isolation & purification; D036103:Nanotechnology / Q000379:methods*; D010100:Oxygen / Q000737:chemistry; D010539:Permeability; D016062:Porosity; D011485:Protein Binding; D058428:Silica Gel; D012822:Silicon Dioxide / Q000737:chemistry*; D017155:Wettability; D014907:Whales / Q000378:metabolism
D020099:Coated Materials, Biocompatible; D006420:Hemeproteins; D006454:Hemoglobins; D008567:Membranes, Artificial; D009211:Myoglobin; D058428:Silica Gel; D012822:Silicon Dioxide; D010100:Oxygen