Deletion of the. Omega. -loop in the active site of staphylococcal nuclease. 2. Effects on protein structure and dynamics
- National Institutes of Health, Bethesda, MD (USA)
- Univ. of Maryland, College Park (USA)
It has been shown that deletion of residues 44-49 from the sequence of staphylococcal nuclease (E43 SNase) results in an enzyme (E43 {Delta}SNase) that is significantly more active than D43 SNase, an enzyme that differs from the wild-type enzyme by deletion of a single methylene group. In addition, both E43 {Delta}SNase and D43 {Delta}SNase are significantly more stable than their respective parent enzymes. Herein they use high-resolution 2D and 3D NMR spectroscopy to characterize the solution conformations of the four enzymes in order to better understand their differences in stability and activity. The backbone assignments of E43 SNase were extended to the three mutant proteins (uniformly {sup 15}N-enriched) by using 2D HSQC, 3D HOHAHA-HMQC, and 3D NOESY-HMQC spectra. The NOE patterns observed for E43 and D43 SNase in solution are consistent with the crystal structures of these proteins. The NOESY data further show that the intact and deleted proteins have essentially the same structures except that (a) the disordered {Omega}-loops, in the intact proteins are replaced by tight type II{prime} turns, formed by residues 43-50-51-52, in the deleted proteins and (b) the orientation of the D43 side chain in crystalline D43 SNase differs from that found for D43 {Delta}SNase in solution. Except for regions neighboring the {Omega}-loops, the intact and deleted proteins show nearly identical amide {sup 15}N and {sup 1}H chemical shifts.
- OSTI ID:
- 5556472
- Journal Information:
- Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 30:15; ISSN 0006-2960; ISSN BICHA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
62 RADIOLOGY AND NUCLEAR MEDICINE
AMIDES
BACTERIA
BARYONS
CHEMICAL SHIFT
ELEMENTARY PARTICLES
ENZYMES
ESTERASES
FERMIONS
HADRONS
HYDROLASES
ISOTOPES
LIGHT NUCLEI
MAGNETIC RESONANCE
MICROORGANISMS
MOLECULAR STRUCTURE
NITROGEN 15
NITROGEN ISOTOPES
NUCLEAR MAGNETIC RESONANCE
NUCLEASES
NUCLEI
NUCLEONS
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OVERHAUSER EFFECT
PHOSPHODIESTERASES
PROTONS
RESONANCE
STABLE ISOTOPES
STAPHYLOCOCCUS