title: Thr-E11 regulates O2 affinity in Cerebratulus lacteus mini-hemoglobin.
abstract: The mini-hemoglobin from Cerebratulus lacteus (CerHb) belongs to a class of globins containing the polar Tyr-B10/Gln-E7 amino acid pair that normally causes low rates of O2 dissociation and ultra-high O2 affinity, which suggest O2 sensing or NO scavenging functions. CerHb, however, has high rates of O2 dissociation (kO2 = 200-600 s(-1)) and moderate O2 affinity (KO2) approximately 1 microm(-1)) as a result of a third polar amino acid in its active site, Thr-E11. When Thr-E11 is replaced by Val, kO2 decreases 1000-fold and KO2 increases 130-fold at pH 7.0, 20 degrees C. The mutation also shifts the stretching frequencies of both heme-bound and photodissociated CO, indicating marked changes of the electrostatic field at the active site. The crystal structure of Thr-E11 --> Val CerHbO2 at 1.70 A resolution is almost identical to that of the wild-type protein (root mean square deviation of 0.12 A). The dramatic functional and spectral effects of the Thr-E11 --> Val mutation are due exclusively to changes in the hydrogen bonding network in the active site. Replacing Thr-E11 with Val "frees" the Tyr-B10 hydroxyl group to rotate toward and donate a strong hydrogen bond to the heme-bound ligand, causing a selective increase in O2 affinity, a decrease of the rate coefficient for O2 dissociation, a 40 cm(-1) decrease in nuCO of heme-bound CO, and an increase in ligand migration toward more remote intermediate sites.
authors.affiliation: Department of Physics-INFM and Center for Excellence in Biomedical Research, University of Genova, Via Dodecaneso 33, 16146 Genova, Italy.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Pesce
authors.forename: Marco
authors.initials: M
authors.lastname: Nardini
authors.forename: Paolo
authors.initials: P
authors.lastname: Ascenzi
authors.forename: Eva
authors.initials: E
authors.lastname: Geuens
authors.forename: Sylvia
authors.initials: S
authors.lastname: Dewilde
authors.forename: Luc
authors.initials: L
authors.lastname: Moens
authors.forename: Martino
authors.initials: M
authors.lastname: Bolognesi
authors.forename: Austen F
authors.initials: AF
authors.lastname: Riggs
authors.forename: Angela
authors.initials: A
authors.lastname: Hale
authors.forename: Pengchi
authors.initials: P
authors.lastname: Deng
authors.forename: G Ulrich
authors.initials: GU
authors.lastname: Nienhaus
authors.forename: John S
authors.initials: JS
authors.lastname: Olson
authors.forename: Karin
authors.initials: K
authors.lastname: Nienhaus
mesh_terms: D000818:Animals; D002248:Carbon Monoxide / Q000378:metabolism; D003460:Crystallization; D006454:Hemoglobins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D007448:Invertebrates / Q000737:chemistry*; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D016296:Mutagenesis; D010100:Oxygen / Q000378:metabolism*; D010782:Photolysis; D017550:Spectroscopy, Fourier Transform Infrared; D055672:Static Electricity; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D013912:Threonine; D014633:Valine
chemical_list: D006454:Hemoglobins; D013912:Threonine; D002248:Carbon Monoxide; D014633:Valine; D010100:Oxygen