Characterization of the Interaction of Photo-Generated Cob(I)inamide with Carbon Dioxide in Anaerobic Aqueous Solution Open Access

Jawdat, BenMaan I. (2010)

Permanent URL:



Characterization of the Interaction of Photo-Generated Cob(I)inamide with
Carbon Dioxide in Anaerobic Aqueous Solution

By BenMaan I. Jawdat

One of the greatest challenges facing today's scientists is the creation of a viable
renewable energy system to meet the demands of a growing population. The use of
solar energy is currently a research focus. The long-term goal of this project is to
develop a light-driven catalytic module in a re-engineered protein, that will be used for
conversion of carbon dioxide (CO2) to a stable fuel in aqueous solution. The selected
protein is the EutB protein from ethanolamine ammonia-lyase, which contains the
native, cobalt-containing cobalamin cofactor. In this project, the reactivity of the free
cobinamide (modified cobalamin) molecule with CO2 is studied in buffered aqueous
solution, outside of the protein environment. Static and time-resolved ultraviolet
(UV)/visible spectroscopy is used to characterize the cobinamide in the absence and
presence of CO2. The fully-reduced cob(I)inamide, was generated by photoreduction of
methylcob(III)inamide with 5'-deazariboflavin/EDTA in anaerobic, buffered (pH=6.0)
solution. The subsequent cob(I)inamide decay reaction was measured by the loss of
absorbance at 387 nm, with and without CO2 in solution. An assay was developed, in
which the photolysis of methylcobalamin, and the subsequent reaction of the
cob(II)alamin photoproduct with oxygen (O2) to form AquoCbl, was used to quantify
the oxygen content in the samples. The careful control of pH and exclusion of oxygen
led to dramatic improvement in the reproducibilty of the cob(I)inamide reaction
measurements, relative to previous attemps. Decay of cob(I)inamide in the absence of
CO2 (τ=9.0 min) was assigned to a proton reduction reaction. Increased time constants
for decay of cob(I)inamide (up to τ=13.5 min) were observed in the presence of CO2.
We propose that the effect is caused by the interaction of CO2 with cob(I)inamide.

Table of Contents

Table of Contents
Chapter Page I. Introduction 1 II. Experimental Procedures 5

- Absorption Spectroscopy


- Photolysis Studies


- Preparation of MeCbi Samples


- Fitting of the Cob(I)inamide Decay Kinetics

8 III. Results

- Absorption Spectra of Different Cobalamin and Cobinamide Redox States

- Assay for the Determination of Oxygen Concentration in the Samples

- Reaction of Cob(I)inamide in Aqueous Solution at pH 6.0

- Reaction of Cob(I)inamide with CO2 in Aqueous Solution at pH 6.0

9 9 11 14 15 IV. Discussion 20

- Establishment of Reliability and Reproducibility in Cobalamin/ Cobinamide - CO2 Reactivity Measurements


- Dihydrogen Formation


- CO2 Binding Interaction with Cob(I)inamide


- Future Experiments and Outlook

23 V. References 25

About this Honors Thesis

Rights statement
  • Permission granted by the author to include this thesis or dissertation in this repository. All rights reserved by the author. Please contact the author for information regarding the reproduction and use of this thesis or dissertation.
  • English
Research field
Committee Chair / Thesis Advisor
Committee Members
Last modified

Primary PDF

Supplemental Files