The solution structure of the histidine‐containing protein (HPr) from Staphylococcus aureus as determined by two‐dimensional ¹H‐NMR spectroscopy

Abstract

The three-dimensional solution structure of the heat-stable phosphocarrier protein HPr from Staphylococcus aureus was determined from two-dimensional NMR data by restrained molecular dynamics. It consists of a large twisted antiparallel P-pleated sheet with four strands A, B, C, and D of amino acids 2-7, 34-37, 40-42 and 60-65. Three right-handed helices A, B, C (amino acids 18-27, 47-53 and 71-85) are positioned on top of this sheet. The aromatic ring of His15 is located in a cleft formed by amino acids 12-17 and 55-58, only the nitrogen (N61) atom which can be phosphorylated by enzyme I is exposed to the water. The side chains of Thrl2 and Argl7 are located close to the histidine ring. The regulatory serine residue (Ser46) is located in a hydrophobic patch, its hydroxyl group is water-accessible but forms hydrogen bonds with the amide groups of the backbone. The general features of the three-dimensional structure are similar to those found in HPr proteins from different microorganisms such as Escherichia coli, Bacillus subtilis and Streptococcus faecalis.

Solution Structure of the IIAChitobiose-HPr Complex of the N,N′-Diacetylchitobiose Branch of the Escherichia coli Phosphotransferase System

“…The structure of the IIA Chb *-HPr complex was largely based on intermolecular NOE data derived from three-dimensional 12 C-filtered/ 13 C-separated three-dimensional NOE experiments in which NOEs are exclusively observed between protons attached to 12 C and protons attached to 13 C. An array of different combinations of isotopically labeled samples, com- prising both uniform and residue-specific labeling (Table 1), was employed to remove any ambiguities in assignment of intermolecular NOEs. An example of the quality of the intermolecular data is provided in Fig.…”

“…3B and 4B), The predominant intermolecular contacts between HPr and IIA Chb * involve helices. The N-terminal halves of helices 1 (residues 16 -27) and 2 (residues 347-348) of HPr interact with the N-terminal halves of helices 1 (residues 18 -33) and 3 (residues 89 and 93) of subunit A of IIA Chb *; while the loop preceding helix 1 (residues [11][12][13][14][15], helix 2 (residues [47][48][49][50][51][52][53], and a stretch of extended strand (residues [55][56][57] of HPr interact with the C-terminal half of helix 2 (residues 58 -73), the loop connecting helices 2 and 3 (residues 76 -82), and the middle half of helix 3 (residues 91-98) of the C subunit of IIA Chb *. The total accessible surface area buried upon complex formation is ϳ1580 Å 2 , comprising ϳ350 Å 2 and ϳ450 Å 2 for subunits A and C, respectively, of IIA Chb *, and ϳ780 Å 2 for HPr (subdivided into ϳ350 and ϳ430 Å 2 for contacts with the A and C subunits of IIA Chb *, respectively).…”

High-Resolution Structure of the Histidine-Containing Phosphocarrier Protein (HPr) fromStaphylococcus aureusand Characterization of Its Interaction with the Bifunctional HPr Kinase/Phosphorylase

“…For HPr from Staphylococcus aureus, only a low-resolution NMR structure, which was solved exclusively with homonuclear methods under a low magnetic field, has been published (29). In this paper, we present a high-resolution structure of HPr from S. aureus as the basis for a study of its interaction with HPrK/P from S. xylosus.…”

“…Simultaneously, the phosphorylation of Ser46 inhibits phosphocarrier activity by perturbing the interaction with phosphorylated EI (3, 48). Furthermore, in some bacteria P-Ser-HPr seems to be involved in additional regulatory mechanisms, called inducer expulsion and inducer exclusion (9,54,61,62).HPr proteins from different microorganisms have been structurally characterized by X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy (10,21,23,24,29,32,41,46,53,60). Although they differ largely in primary structure, their general folding structure is well conserved.…”

“…HPr proteins from different microorganisms have been structurally characterized by X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy (10,21,23,24,29,32,41,46,53,60). Although they differ largely in primary structure, their general folding structure is well conserved.…”