Low frequency dynamics of Cystathionine beta-synthase

Abstract

Femtosecond coherence spectroscopy is used to study the low frequency dynamics of cystathionine beta-synthase (CBS). CBS is a pyridoxal-5′-phosphate-dependent heme enzyme with cysteine and histidine axial ligands that catalyzes the condensation of serine and homocysteine to form cystathionine. Resonance excitation near the maximum of the ferric state Soret band reveals a mode near ∼40 cm−1 (phase ∼pi/2). The phase indicates that the initial non-equilibrium coherent wavepacket for this mode is dominated by a momentum displacement. This is consistent with doming of the ferric five-coordinate species and suggests photolysis of the histidine ligand. When exciting on the red side of the Soret band, a mode near ∼25 cm−1 is observed that exhibits a phase jump of ∼pi for blue-side excitation. This mode may involve the response of an unphotolyzed fraction of hot ferric six-coordinate species, subsequent to ultrafast non-radiative decay. A strong correlation between the “detuned” coherence spectrum (which reveals higher frequencies) and the Raman spectrum is also demonstrated. Normal coordinate structural decomposition of the ferric heme crystal structure predicts strong saddling, doming, and ruffling modes and they are observed in the coherence spectra. The relative intensities of these modes are monitored as a function of pH in order to explore the potential correlation between redox equilibria, pH, and protein-induced heme structural perturbations. The low frequency spectra of ferrous CBS and its NO-bound complex were also obtained, along with the CO rebinding kinetics. The geminate rebinding of CO to CBS was found to be unusually fast and similar to that of CooA.