Irondistances inhemoglobin: Comparison ofx-ray crystallographic andextended x-ray absorption finestructure studies

A comparison ispresented ofthestructures obtained around theiron atomindeoxyhemoglobin (Hb). The datacomefromextended x-rayabsorption finestructure (EXAFS) studies oftheironwhich gaveFe-porphyrin nitrogen distances of2.06± 0.01A,andfromthemostrecent high- resolution x-ray crystallographic study, which gaveexactly the samedistance-2.06 ± 0.02A.Thedistance ofFeabove the plane oftheporphyrin nitrogens was0.38 ± 0.04Afromthe crystallographic study; this value isnotfarfromtheupper limit ofthedistances 0.20± °:10 A calculated fromtheEXAFS experiment bytriangulation. These distances above thenitro- genplane areshorter thanthose estimated intheearliest x-ray structures. A recent note(1)byoneofus(R.G.S.) tookissue with Galloway's (2)criticism oftheearly extended x-ray absorp- tionfinestructure (EXAFS)studies ofdeoxyhemoglobin (Hb)byEisenberger etal. (3). TheseEXAFSmeasurements hadobtained Fe-porphyrin nitrogen (Np)distances of2.055 ±0.01 A,which hadbeensupported bysubsequent EXAFS studies byPerutz etal.(4), inwhichFe-Npequaled 2.06+ 0.01A.Eisenberger etal.(3)intheir early workcalculated Fe-Ct, thedistance fromFetothecenter oftheNpplane, by taking theCt-Npdistance tobe2.04 A(from x-ray structures ofmodel compounds andresonance Ramanmeasurements of Hb)andtaking thehypotenuse (the Fe-Npdistance) tobe 2.06 A asmeasured. Including estimates oftheir errors, they calculated thattheFe-Ctdistance was0.20± 8:28 A.The Fe-Ctdistance soderived wascompared withresults of earlier moderate-resolution (2.5 Aandlower) x-ray studies of Hb(5,6)withthesuggestion that thelatter wereinerror. We areconcerned that thenotemayhavegiven theimpression thatEXAFSandprotein x-raycrystallography presently yield different results inHb.Infact, thetwomethods presently yield precisely thesameresults, aswasshownby ahigh-resolution (1.74 A)x-ray crystallographic study of Hb (7)towhichR.G.S.didnotrefer inhisanswerto Galloway's criticism. Thehigh-resolution x-ray study ofHb(7)yielded aFe-Np distance of2.06 ± 0.02A,inexact agreement withthevalue 2.06 ±0.01 AfromEXAFS(3, 4). Thex-ray study (7) yielded aFe-Ctdistance of0.38± 0.04A,ingoodagreement with thevalues 0.418A and0.399A intwoofthethree five- coordinated Fe2+modelcompounds listed inref. 1 but slightly larger thanthevalue 0.28A inthethird. Thevalues Fe-Np= 2.06A andFe-Ct= 0.38A yield anestimate for Ct-Npof2.02 A,which isabout 0.01 Aless thantheshortest Fe-Ctdistance amongthethree modelcompounds and0.02 Aless thanthevalue usedintriangulating theEXAFSresults (3). TheEXAFSandhigh-resolution x-ray results arecom- pared inTable 1.Goodagreement isseenbetween thetwo