DiGeorge critical region 8 (DGCR8) is a double-cysteine-ligated heme protein.

Authors

Ian Barr, University of California, Los AngelesFollow
Aaron T. Smith, University of California, Los AngelesFollow
Rachel Senturia, University of California, Los AngelesFollow
Yanqiu Chen, University of California, Los Angeles
Brooke D. Scheidemantle, University of California, Los AngelesFollow
Judith N. Burstyn, University of California, Los AngelesFollow
Feng Guo, University of California, Los AngelesFollow

Department

Natural Sciences and Mathematics

Document Type

Article

Source

Journal of Biological Chemistry

Publication Date

5-13-2011

ISSN

1083-351X

Volume

286

Issue

19

First Page

16716 - 16725

Abstract

All known heme-thiolate proteins ligate the heme iron using one cysteine side chain. We previously found that DiGeorge Critical Region 8 (DGCR8), an essential microRNA processing factor, associates with heme of unknown redox state when overexpressed in Escherichia coli. On the basis of the similarity of the 450-nm Soret absorption peak of the DGCR8-heme complex to that of cytochrome P450 containing ferrous heme with CO bound, we identified cysteine 352 as a probable axial ligand in DGCR8. Here we further characterize the DGCR8-heme interaction using biochemical and spectroscopic methods. The DGCR8-heme complex is highly stable, with a half-life exceeding 4 days. Mutation of the conserved proline 351 to an alanine increases the rate of heme dissociation and allows the DGCR8-heme complex to be reconstituted biochemically. Surprisingly, DGCR8 binds ferric heme without CO to generate a hyperporphyrin spectrum. The electronic absorption, magnetic circular dichroism, and electron paramagnetic resonance spectra of the DGCR8-heme complex suggest a ferric heme bearing two cysteine ligands. This model was further confirmed using selenomethionine-substituted DGCR8 and mercury titration. DGCR8 is the first example of a heme-binding protein with two endogenous cysteine side chains serving as axial ligands. We further show that native DGCR8 binds heme when expressed in eukaryotic cells. This study provides a chemical basis for understanding the function of the DGCR8-heme interaction in microRNA maturation.

PubMed ID

21454614

Creative Commons License

This work is licensed under a Creative Commons Attribution-NonCommercial-No Derivative Works 4.0 International License.