Published online 29 March 2017 Nucleic Acids Research, 2017, Vol. 45, No. 10 6217–6227 doi: 10.1093/nar/gkx222 Structure of acid deoxyribonuclease Armando Varela-Ramirez1, Jan Abendroth2,3, Adrian A. Mejia1, Isabelle Q. Phan2,4, Donald D. Lorimer2,3, Thomas E. Edwards2,3,* and Renato J. Aguilera1,*
1Department of Biological Sciences, Border Biomedical Research Center, The University of Texas at El Paso, El Paso, TX 79968, USA, 2Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, WA 98110, USA, 3Beryllium Discovery Corp., 7869 NE Day Road West, Bainbridge Island, WA 98110, USA and 4Center for Infectious Disease Research (formerly Seattle Biomedical Research Institute), 307 Westlake Ave N, Seattle, WA 98109, USA
Received February 16, 2017; Revised March 20, 2017; Editorial Decision March 22, 2017; Accepted March 23, 2017
ABSTRACT of human DNase I has been reported recently (6), no crys- tal structure has been reported for any DNase II. Deoxyribonuclease II (DNase II) is also known as acid DNase II belongs to a unique family of nucleases that deoxyribonuclease because it has optimal activity is present in most eukaryotes. Although DNase II enzymes at the low pH environment of lysosomes where it are widely distributed in mammalian tissues (7), they are is typically found in higher eukaryotes. Interestingly, most commonly found within lysosomes or phagolyso- DNase II has also been identified in a few genera of somes (8). The acidic pH found within the lysosome or- bacteria and is believed to have arisen via horizon- ganelle makes it the perfect environment for the optimum tal transfer. Here, we demonstrate that recombinant enzymatic activity of DNase II, which is also referred to as acid deoxyribonuclease for this reason. Furthermore, gen- Burkholderia thailandensis DNase II is highly active at low pH in the absence of divalent metal ions, simi- eration of the 3 -phosphorylated and 5 -hydroxyl ends has lar to eukaryotic DNase II. The crystal structure of B. been predicted to prevent ready integration of foreign DNA into the host genome since repair enzymes typically rec- thailandensis DNase II shows a dimeric quaternary ognize 5 -phosphorylated and 3 -hydroxyl substrates (9,10). structure which appears capable of binding double- Although DNase II has been shown to have preference for stranded DNA. Each monomer of B. thailandensis runs of purines, the structure of DNA appears to be more DNase II exhibits a similar overall fold as phospho- important than the actual sequence for cleavage (11,12). lipase D (PLD), phosphatidylserine synthase (PSS) This was elegantly demonstrated by early experiments in and tyrosyl-DNA phosphodiesterase (TDP), and con- which the cleavage pattern of DNase II was altered in the served catalytic residues imply a similar mechanism. presence of DMSO, which significantly affects the structure The structural and biochemical data presented here of DNA (11). It has also been demonstrated that DNase II provide insights into the atomic structure and cat- cleaves single stranded DNA and can cleave stronger on one alytic mechanism of DNase II. strand (due to presence of purines) than the other (13). Due to its cleavage preference, it was previously postulated that DNase II might have a narrow binding domain that could bind one strand with minimal interference from the other INTRODUCTION strand (13). In humans and other vertebrates, DNase II appears to Deoxyribonuclease (DNase) enzymes perform a variety of have undergone a gene duplication event resulting in two important cellular roles by degrading DNA via hydroly- closely related genes dnase IIα and dnase IIβ. DNase IIα is sis of its phosphodiester backbone. Deoxyribonuclease I expressed in all tissues and mutation of DNase IIα in mice (DNase I) enzymes cleave single or double-stranded DNA results in an embryonic lethal phenotype as this enzyme and require divalent metal ions to hydrolyze DNA yield- plays a critical role in phagocyte-mediated apoptotic DNA ing 3 -hydroxyl and 5 -phosphorylated products. In con- degradation (14,15). DNase IIβ is also known as DNase II trast, deoxyribonuclease II (DNase II) hydrolyzes DNA at like acid DNase (DLAD) (4) and is expressed in only a few low pH to yield 3 -phosphorylated and 5 -hydroxyl prod- tissues such as the eye lens. It has been shown to have an ucts (Figure 1A) (1). Strikingly, unlike many phosphoryl essential function in clearing DNA from the lens, and dnase transfer enzymes such as other nucleases (2) and all nucleic IIβ deficient mice readily develop cataracts (16,17). Apart acid polymerases (3), DNase II does not require divalent from its obvious role in DNA degradation, DNase II was metal ions for catalysis (4,5). Although a crystal structure
*To whom correspondence should be addressed. Tel: +1 206 780 8949; Email: [email protected] Correspondence may also be addressed to Renato Aguilera. Tel: +1 915 747 6852; Email: [email protected]