Funneled angle landscapes for helical proteins

We use crystallographic data for four helical iron proteins (cytochrome c-b₅₆₂, cytochrome c′, sperm whale myoglobin, human cytoglobin) to calculate radial and angular signatures as each unfolds from the native state stepwise though four unfolded states. From these data we construct an angle phase diagram to display the evolution of each protein from its native state; and, in turn, the phase diagram is used to construct a funneled angle landscape for comparison with the topography of its folding energy landscape. We quantify the departure of individual helical and turning regions from the areal, angular profile of corresponding regions of the native state. This procedure allows us to identify the similarities and differences among individual helical and turning regions in the early stages of unfolding of the four helical heme proteins

Structural stability of the SARS-CoV-2 main protease: Can metal ions affect function?

We have investigated the structural stability of the SARS (Severe acute respiratory syndrome)-CoV-2 main protease monomer (Mpro). We quantified the spatial and angular changes in the structure using two independent analyses, one based on a spatial metrics (δ, ratio), the second on angular metrics. The order of unfolding of the 10 helices in Mpro is characterized by beta vs alpha plots similar to those of cytochromes and globins. The longest turning region is anomalous in the earliest stage of unfolding. In an investigation of excluded-volume effects, we found that the maximum spread in average molecular-volume values for Mpro, cytochrome c-b₅₆₂, cytochrome c’, myoglobin, and cytoglobin is ~10 ų. This apparent universality is a consequence of the dominant contributions from six residues: ALA, ASP, GLU, LEU, LYS and VAL. Of the seven Mpro histidines, residues 41, 163, 164, and 246 are in stable H-bonded regions; metal ion binding to one or more of these residues could break up the H-bond network, thereby affecting protease function. Our analysis also indicated that metal binding to cysteine residues 44 and 145 could disable the enzyme

Unfolding cytochromes c-b₅₆₂ and Rd apo b₅₆₂

We have analyzed the early stages of unfolding of cytochromes c-b₅₆₂ (PDB ID: 2BC5) and Rd apo b₅₆₂ (PDB ID: 1YYJ). Our geometrical approach proceeds from an analysis of the crystal structure reported for each protein. We quantify, residue-by-residue and region-by-region, the spatial and angular changes in the structure as the protein denatures, and quantify differences that result from the seven residues that differ in the two proteins. Using two independent analyses, one based on spatial metrics and the second on angular metrics, we establish the order of unfolding of the five helices in cyt c-b₅₆₂ and the four helices in the apo protein. For the two helices nearest the N-terminal end of both proteins, the ones in the apo protein unfold first. For the two helices nearest the C-terminal end, the interior helix of the apo protein unfolds first, whereas the terminal helix of the holo protein unfolds first. Excluded-volume effects (repulsive interactions) are minimized in turning regions; the overall range in Δ values is Δ = 36.3 ų for cyt c-b₅₆₂ and Δ = 36.6 ų for the apo protein, whereas the span for all 20 amino acids is Δ = 167.7 ų. As our work indicates that the interior helix of cytochrome c-b₅₆₂ is the first to fold, we suggest that this helix protects the heme from misligation, consistent with ultrafast folding over a minimally frustrated funneled landscape

Structural stability of the SARS-CoV-2 main protease: Can metal ions affect function?

We have investigated the structural stability of the SARS (Severe acute respiratory syndrome)-CoV-2 main protease monomer (Mpro). We quantified the spatial and angular changes in the structure using two independent analyses, one based on a spatial metrics (δ, ratio), the second on angular metrics. The order of unfolding of the 10 helices in Mpro is characterized by beta vs alpha plots similar to those of cytochromes and globins. The longest turning region is anomalous in the earliest stage of unfolding. In an investigation of excluded-volume effects, we found that the maximum spread in average molecular-volume values for Mpro, cytochrome c-b₅₆₂, cytochrome c’, myoglobin, and cytoglobin is ~10 ų. This apparent universality is a consequence of the dominant contributions from six residues: ALA, ASP, GLU, LEU, LYS and VAL. Of the seven Mpro histidines, residues 41, 163, 164, and 246 are in stable H-bonded regions; metal ion binding to one or more of these residues could break up the H-bond network, thereby affecting protease function. Our analysis also indicated that metal binding to cysteine residues 44 and 145 could disable the enzyme

Unfolding cytochromes c-b₅₆₂ and Rd apo b₅₆₂

We have analyzed the early stages of unfolding of cytochromes c-b₅₆₂ (PDB ID: 2BC5) and Rd apo b₅₆₂ (PDB ID: 1YYJ). Our geometrical approach proceeds from an analysis of the crystal structure reported for each protein. We quantify, residue-by-residue and region-by-region, the spatial and angular changes in the structure as the protein denatures, and quantify differences that result from the seven residues that differ in the two proteins. Using two independent analyses, one based on spatial metrics and the second on angular metrics, we establish the order of unfolding of the five helices in cyt c-b₅₆₂ and the four helices in the apo protein. For the two helices nearest the N-terminal end of both proteins, the ones in the apo protein unfold first. For the two helices nearest the C-terminal end, the interior helix of the apo protein unfolds first, whereas the terminal helix of the holo protein unfolds first. Excluded-volume effects (repulsive interactions) are minimized in turning regions; the overall range in Δ values is Δ = 36.3 ų for cyt c-b₅₆₂ and Δ = 36.6 ų for the apo protein, whereas the span for all 20 amino acids is Δ = 167.7 ų. As our work indicates that the interior helix of cytochrome c-b₅₆₂ is the first to fold, we suggest that this helix protects the heme from misligation, consistent with ultrafast folding over a minimally frustrated funneled landscape

Structural Stability of Intelectin-1

We study the structural stability of helical and non-helical regions in chain A of human intelectin-1. Using a geometrical model introduced previously, a computational analysis based on the recently reported crystal structure of this protein by Kiessling et al. [Nature Struct. Mole. Bio. 22 (2015), 603] is carried out to quantify the resiliency of the native state to steric perturbations. Response to these perturbations is characterized by calculating, relative to the native state, the lateral, radial and angular displacements of n-residue segments of the polypeptide chain centered on each residue. By quantifying the stability of the protein through six stages of unfolding, we are able to identify regions in chain A of intelectin-1 which are markedly affected by structural perturbations versus those which are relatively unaffected, the latter suggesting that the native-state geometry of these regions is essentially conserved. Importantly, residues in the vicinity of calcium ions comprise a conserved region, suggesting that Ca ions play a role not only in the coordination of carbohydrate hydroxyl groups, but in preserving the integrity of the structure

Relaxation of structural constraints during Amicyanin unfolding

We study the thermal unfolding of amicyanin by quantifying the resiliency of the native state to structural perturbations. Three signatures characterizing stages of unfolding are identified. The first signature, lateral extension of the polypeptide chain, is calculated directly from the reported crystallographic data. Two other signatures, the radial displacement of each residue from Cu(II) and the angular spread in the chain as the protein unfolds, are calculated using crystallographic data in concert with a geometrical model we introduced previously (J.J. Kozak, H. B. Gray, R. A. Garza-López, J. Inorg. Biochem. 155(2016) 44–55). Particular attention is paid to the resiliency of the two beta sheets in amicyanin. The resiliency of residues in the near neighborhood of the Cu center to destabilization provides information on the persistence of the entatic state. Similarly, examining the resiliency of residues intercalated between structured regions (beta sheets, the alpha helix) provides a basis for identifying a “hydrophobic core.” A principal focus of our study is to compare results obtained using our geometrical model with the experimental results (C. La Rosa, D. Milardi, D. M. Grasso, M. P. Verbeet, G. W. Canters, L. Sportelli, R. Guzzi, Eur. Biophy. J.30(8),(2002) 559–570) on the denaturation of amicyanin, and we show that our results support a classical model proposed by these authors

Structural Stability of Intelectin-1

We study the structural stability of helical and non-helical regions in chain A of human intelectin-1. Using a geometrical model introduced previously, a computational analysis based on the recently reported crystal structure of this protein by Kiessling et al. [Nature Struct. Mole. Bio. 22 (2015), 603] is carried out to quantify the resiliency of the native state to steric perturbations. Response to these perturbations is characterized by calculating, relative to the native state, the lateral, radial and angular displacements of n-residue segments of the polypeptide chain centered on each residue. By quantifying the stability of the protein through six stages of unfolding, we are able to identify regions in chain A of intelectin-1 which are markedly affected by structural perturbations versus those which are relatively unaffected, the latter suggesting that the native-state geometry of these regions is essentially conserved. Importantly, residues in the vicinity of calcium ions comprise a conserved region, suggesting that Ca ions play a role not only in the coordination of carbohydrate hydroxyl groups, but in preserving the integrity of the structure