93 research outputs found
Molecular basis of RNA guanine-7 methyltransferase (RNMT) activation by RAM
Maturation and translation of mRNA in eukaryotes requires the addition of the 7-methylguanosine cap. In vertebrates, the cap methyltransferase, RNA guanine-7 methyltransferase (RNMT), has an activating subunit, RNMT-Activating Miniprotein (RAM). Here we report the first crystal structure of the human RNMT in complex with the activation domain of RAM. A relatively unstructured and negatively charged RAM binds to a positively charged surface groove on RNMT, distal to the active site. This results in stabilisation of a RNMT lobe structure which co-evolved with RAM and is required for RAM binding. Structure-guided mutagenesis and molecular dynamics simulations reveal that RAM stabilises the structure and positioning of the RNMT lobe and the adjacent α-helix hinge, resulting in optimal positioning of helix A which contacts substrates in the active site. Using biophysical and biochemical approaches, we observe that RAM increases the recruitment of the methyl donor, AdoMet (S-adenosyl methionine), to RNMT. Thus we report the mechanism by which RAM allosterically activates RNMT, allowing it to function as a molecular rheostat for mRNA cap methylation
Electronic Coherence Dephasing in Excitonic Molecular Complexes: Role of Markov and Secular Approximations
We compare four different types of equations of motion for reduced density
matrix of a system of molecular excitons interacting with thermodynamic bath.
All four equations are of second order in the linear system-bath interaction
Hamiltonian, with different approximations applied in their derivation. In
particular we compare time-nonlocal equations obtained from so-called
Nakajima-Zwanzig identity and the time-local equations resulting from the
partial ordering prescription of the cummulant expansion. In each of these
equations we alternatively apply secular approximation to decouple population
and coherence dynamics from each other. We focus on the dynamics of intraband
electronic coherences of the excitonic system which can be traced by coherent
two-dimensional spectroscopy. We discuss the applicability of the four
relaxation theories to simulations of population and coherence dynamics, and
identify features of the two-dimensional coherent spectrum that allow us to
distinguish time-nonlocal effects.Comment: 14 pages, 8 figure
Full characterization of vibrational coherence in a porphyrin chromophore by two-dimensional electronic spectroscopy
In this work we present experimental and calculated two-dimensional electronic spectra for a 5,15-bisalkynyl porphyrin chromophore. The lowest energy electronic Qy transition couples mainly to a single 380 cm–1 vibrational mode. The two-dimensional electronic spectra reveal diagonal and cross peaks which oscillate as a function of population time. We analyze both the amplitude and phase distribution of this main vibronic transition as a function of excitation and detection frequencies. Even though Feynman diagrams provide a good indication of where the amplitude of the oscillating components are located in the excitation-detection plane, other factors also affect this distribution. Specifically, the oscillation corresponding to each Feynman diagram is expected to have a phase that is a function of excitation and detection frequencies. Therefore, the overall phase of the experimentally observed oscillation will reflect this phase dependence. Another consequence is that the overall oscillation amplitude can show interference patterns resulting from overlapping contributions from neighboring Feynman diagrams. These observations are consistently reproduced through simulations based on third order perturbation theory coupled to a spectral density described by a Brownian oscillator model
A two-lane mechanism for selective biological ammonium transport
The transport of charged molecules across biological membranes faces the dual problem of accommodating charges in a highly hydrophobic environment while maintaining selective substrate translocation. This has been the subject of a particular controversy for the exchange of ammonium across cellular membranes, an essential process in all domains of life. Ammonium transport is mediated by the ubiquitous Amt/Mep/Rh transporters that includes the human Rhesus factors. Here, using a combination of electrophysiology, yeast functional complementation and extended molecular dynamics simulations, we reveal a unique two-lane pathway for electrogenic NH4+ transport in two archetypal members of the family, the transporters AmtB from Escherichia coli and Rh50 from Nitrosomonas europaea. The pathway underpins a mechanism by which charged H+ and neutral NH3 are carried separately across the membrane after NH4+ deprotonation. This mechanism defines a new principle of achieving transport selectivity against competing ions in a biological transport process
Capturing the essence of folding and functions of biomolecules using Coarse-Grained Models
The distances over which biological molecules and their complexes can
function range from a few nanometres, in the case of folded structures, to
millimetres, for example during chromosome organization. Describing phenomena
that cover such diverse length, and also time scales, requires models that
capture the underlying physics for the particular length scale of interest.
Theoretical ideas, in particular, concepts from polymer physics, have guided
the development of coarse-grained models to study folding of DNA, RNA, and
proteins. More recently, such models and their variants have been applied to
the functions of biological nanomachines. Simulations using coarse-grained
models are now poised to address a wide range of problems in biology.Comment: 37 pages, 8 figure
Excitation Dynamics and Relaxation in a Molecular Heterodimer
The exciton dynamics in a molecular heterodimer is studied as a function of
differences in excitation and reorganization energies, asymmetry in transition
dipole moments and excited state lifetimes. The heterodimer is composed of two
molecules modeled as two-level systems coupled by the resonance interaction.
The system-bath coupling is taken into account as a modulating factor of the
energy gap of the molecular excitation, while the relaxation to the ground
state is treated phenomenologically. Comparison of the description of the
excitation dynamics modeled using either the Redfield equations (secular and
full forms) or the Hierarchical quantum master equation (HQME) is demonstrated
and discussed. Possible role of the dimer as an excitation quenching center in
photosynthesis self-regulation is discussed. It is concluded that the
system-bath interaction rather than the excitonic effect determines the
excitation quenching ability of such a dimer
Role of Active Site Rigidity in Activity: MD Simulation and Fluorescence Study on a Lipase Mutant
Relationship between stability and activity of enzymes is maintained by underlying conformational flexibility. In thermophilic enzymes, a decrease in flexibility causes low enzyme activity while in less stable proteins such as mesophiles and psychrophiles, an increase in flexibility is associated with enhanced enzyme activity. Recently, we identified a mutant of a lipase whose stability and activity were enhanced simultaneously. In this work, we probed the conformational dynamics of the mutant and the wild type lipase, particularly flexibility of their active site using molecular dynamic simulations and time-resolved fluorescence techniques. In contrast to the earlier observations, our data show that active site of the mutant is more rigid than wild type enzyme. Further investigation suggests that this lipase needs minimal reorganization/flexibility of active site residues during its catalytic cycle. Molecular dynamic simulations suggest that catalytically competent active site geometry of the mutant is relatively more preserved than wild type lipase, which might have led to its higher enzyme activity. Our study implies that widely accepted positive correlation between conformation flexibility and enzyme activity need not be stringent and draws attention to the possibility that high enzyme activity can still be accomplished in a rigid active site and stable protein structures. This finding has a significant implication towards better understanding of involvement of dynamic motions in enzyme catalysis and enzyme engineering through mutations in active site
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