Abstract:Understanding the role of stability strengths and weaknesses in proteins is a key objective for rationalizing their dynamical and functional properties such as conformational changes, catalytic activity, and protein-protein and protein-ligand interactions. We present BRANEart, a new, fast and accurate method to evaluate the per-residue contributions to the overall stability of membrane proteins. It is based on an extended set of recently introduced statistical potentials derived from membrane protein structure… Show more
“…2 A). Figure 2 PyMol 40 depictions of GF19 compared to the LL37 parent molecule and its other fragments. ( A ) The designed, full-length peptide structures with their accessible surface area (ASA) are shown with a coloring scheme based on atom types (N: blue, O: red).…”
Section: Resultsmentioning
confidence: 99%
“…A second measure, the Membrane Propensity index (or MPr score) 40 , was implemented as a cross-validation method in a complementary approach to that of rGb . MPr describes the membrane preference of a protein unit which can, in turn, be redirected to indirectly interpret the aqueous preference of the globular protein units.…”
Section: Resultsmentioning
confidence: 99%
“…GF17 and KR12 had similar degrees of instability, while GF19 was slightly less unstable in an aqueous environment. Distribution of the inhomogeneous instabilities locally on the peptide structures could be seen from the comparative visual portray of the BRANEart 40 MPr map on LL37 and the designed peptide structures (Fig. 2 B).…”
Corneal HSV-1 infections are a leading cause of infectious blindness globally by triggering tissue damage due to the intense inflammation. HSV-1 infections are treated mainly with antiviral drugs that clear the infections but are inefficient as prophylactics. The body produces innate cationic host defence peptides (cHDP), such as the cathelicidin LL37. Various epithelia, including the corneal epithelium, express LL37. cHDPs can cause disintegration of pathogen membranes, stimulate chemokine production, and attract immune cells. Here, we selected GF17, a peptide containing the LL37 fragment with bioactivity but with minimal cytotoxicity, and added two cell-penetrating amino acids to enhance its activity. The resulting GF19 was relatively cell-friendly, inducing only partial activation of antigen presenting immune cells in vitro. We showed that HSV-1 spreads by tunneling nanotubes in cultured human corneal epithelial cells. GF19 given before infection was able to block infection, most likely by blocking viral entry. When cells were sequentially exposed to viruses and GF19, the infection was attenuated but not arrested, supporting the contention that the GF19 mode of action was to block viral entry. Encapsulation into silica nanoparticles allowed a more sustained release of GF19, enhancing its activity. GF19 is most likely suitable as a prevention rather than a virucidal treatment.
“…2 A). Figure 2 PyMol 40 depictions of GF19 compared to the LL37 parent molecule and its other fragments. ( A ) The designed, full-length peptide structures with their accessible surface area (ASA) are shown with a coloring scheme based on atom types (N: blue, O: red).…”
Section: Resultsmentioning
confidence: 99%
“…A second measure, the Membrane Propensity index (or MPr score) 40 , was implemented as a cross-validation method in a complementary approach to that of rGb . MPr describes the membrane preference of a protein unit which can, in turn, be redirected to indirectly interpret the aqueous preference of the globular protein units.…”
Section: Resultsmentioning
confidence: 99%
“…GF17 and KR12 had similar degrees of instability, while GF19 was slightly less unstable in an aqueous environment. Distribution of the inhomogeneous instabilities locally on the peptide structures could be seen from the comparative visual portray of the BRANEart 40 MPr map on LL37 and the designed peptide structures (Fig. 2 B).…”
Corneal HSV-1 infections are a leading cause of infectious blindness globally by triggering tissue damage due to the intense inflammation. HSV-1 infections are treated mainly with antiviral drugs that clear the infections but are inefficient as prophylactics. The body produces innate cationic host defence peptides (cHDP), such as the cathelicidin LL37. Various epithelia, including the corneal epithelium, express LL37. cHDPs can cause disintegration of pathogen membranes, stimulate chemokine production, and attract immune cells. Here, we selected GF17, a peptide containing the LL37 fragment with bioactivity but with minimal cytotoxicity, and added two cell-penetrating amino acids to enhance its activity. The resulting GF19 was relatively cell-friendly, inducing only partial activation of antigen presenting immune cells in vitro. We showed that HSV-1 spreads by tunneling nanotubes in cultured human corneal epithelial cells. GF19 given before infection was able to block infection, most likely by blocking viral entry. When cells were sequentially exposed to viruses and GF19, the infection was attenuated but not arrested, supporting the contention that the GF19 mode of action was to block viral entry. Encapsulation into silica nanoparticles allowed a more sustained release of GF19, enhancing its activity. GF19 is most likely suitable as a prevention rather than a virucidal treatment.
“…With our conventions, negative Δ G i values identify regions called stability strengths, which strongly contribute to the stability of the overall protein structure. Positive Δ G i values, however, indicate stability weaknesses, which correspond to regions that are not stable by themselves but are often optimized for functional reasons rather than for their contribution to the stability of the native structure. , …”
Section: Methodsmentioning
confidence: 99%
“…Positive ΔG i values, however, indicate stability weaknesses, which correspond to regions that are not stable by themselves but are often optimized for functional reasons rather than for their contribution to the stability of the native structure. 20,29 ■…”
A general limitation of the use of enzymes in biotechnological
processes under sometimes nonphysiological conditions is the complex
interplay between two key quantities, enzyme activity and stability,
where the increase of one is often associated with the decrease of
the other. A precise stability-activity trade-off is necessary for
the enzymes to be fully functional, but its weight in different protein
regions and its dependence on environmental conditions is not yet
elucidated. To advance this issue, we used the formalism that we have
recently developed to effectively identify stability strength and
weakness regions in protein structures and applied it to a large set
of globular enzymes with known experimental structure and catalytic
sites. Our analysis showed a striking oscillatory pattern of free
energy compensation centered on the catalytic region. Indeed, catalytic
residues are usually nonoptimal with respect to stability, but residues
in the first shell around the catalytic site are, on the average,
stability strengths and thus compensate for this lack of stability;
residues in the second shell are weaker again, and so on. This trend
is consistent across all enzyme families. It is accompanied by a similar,
but less pronounced, pattern of residue conservation across evolution.
In addition, we analyzed cold- and heat-adapted enzymes separately
and highlighted different patterns of stability strengths and weaknesses,
which provide insight into the longstanding problem of catalytic rate
enhancement in cold environments. The successful comparison of our
stability and conservation results with experimental fitness data,
obtained by deep mutagenesis scanning, led us to propose criteria
for improving catalytic activity while maintaining enzyme stability,
a key goal in enzyme design.
A general limitation of the use of enzymes in biotechnological processes under sometimes non-physiological conditions is the complex interplay between two key quantities, enzyme activity and stability, where the increase of one is often associated with the decrease of the other. A precise stability-activity trade-off is necessary for the enzymes to be fully functional, but its weight in different protein regions and its dependence on environmental conditions is not yet elucidated. To advance this issue, we used the formalism that we have recently developed to effectively identify stability strength and weakness regions in protein structures, and applied it to a large set of globular enzymes with known experimental structure and catalytic sites. Our analysis showed a striking oscillatory pattern of free energy compensation centered on the catalytic region. Indeed, catalytic residues are usually non-optimal with respect to stability, but residues in the first shell around the catalytic site are, on the average, stability strengths and thus compensate for this lack of stability; residues in the second shell are weaker again, and so on. This trend is consistent across all enzyme families. It is accompanied by a similar, but less pronounced, pattern of residue conservation across evolution. In addition, we analyzed cold- and heat-adapted enzymes separately and highlighted different patterns of stability strengths and weaknesses, which provide insight into the longstanding problem of catalytic rate enhancement in cold environments. The successful comparison of our stability and conservation results with experimental fitness data, obtained by deep mutagenesis scanning, led us to propose criteria for improving catalytic activity while maintaining enzyme stability, a key goal in enzyme design.
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