Distinct heme active-site structure in lactoperoxidase revealed by resonance Raman spectroscopy.
Low-frequency resonance Raman spectra of the cyanide and carbon monoxide adducts of lactoperoxidase are obtained with Soret excitation. The nu(Fe-CN) and delta(Fe-C-N) modes are detected at 360 and 453 cm-1, respectively. Upon the isotopic substitution of 13C14N, 12C15N, and 13C15N, the band at 453 cm-1 in the natural abundance adduct shifts to 448, 452, and 445 cm-1, while the 360-cm-1 peak shifts to 358, 357, and 356 cm-1, respectively. The 360-cm-1 band is shifted to 355 cm-1 when the pH is changed from 7.0 to 10.5. On the basis of a previous normal-mode analysis of the cyanoferric adduct of myeloperoxidase, a bent Fe-C-N linkage is suggested for the cyanide adduct of lactoperoxidase. The nu(Fe-CN) (374 cm-1) and delta(Fe-C-N) (480 cm-1) modes are observed for the cyanide adduct of reduced lactoperoxidase. For the carbon monoxide adduct, the nu(Fe-CO) (533 cm-1) and delta(Fe-C-O) (578 cm-1) modes at pH 7.0 are observed to shift to 498 and 570 cm-1 as the pH is raised from 7.0 to 10.0. The strong intensity of delta(Fe-C-O) at both acid and alkaline pHs, along with a suggested bent structure of the Fe-C-N moiety, implies a narrow heme pocket for lactoperoxidase.
Chemistry Department, Marquette University, Milwaukee, Wisconsin 53233.
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Hu
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Treat
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Kincaid
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D003486:Cyanides / Q000378:metabolism / Q000494:pharmacology; D005260:Female; D006418:Heme; D007784:Lactoperoxidase / Q000737:chemistry* / Q000302:isolation & purification; D008892:Milk / Q000201:enzymology; D011487:Protein Conformation; D013059:Spectrum Analysis, Raman / Q000379:methods
D003486:Cyanides; D006418:Heme; D007784:Lactoperoxidase