The enzyme uracil DNA glycosylase (UNG) excises unwanted uracil bases in the genome using an extrahelical base recognition mechanism. Efficient removal of uracil is essential for prevention of C-to-T transition mutations arising from cytosine deamination, cytotoxic U*A pairs arising from incorporation of dUTP in DNA, and for increasing immunoglobulin gene diversity during the acquired immune response. A central event in all of these UNG-mediated processes is the singling out of rare U*A or U*G base pairs in a background of approximately 10(9) T*A or C*G base pairs in the human genome. Here we establish for the human and Escherichia coli enzymes that discrimination of thymine and uracil is initiated by thermally induced opening of T*A and U*A base pairs and not by active participation of the enzyme. Thus, base-pair dynamics has a critical role in the genome-wide search for uracil, and may be involved in initial damage recognition by other DNA repair glycosylases.

Background-Clinical and radiologic predictors of cerebral infarction occurrence and location after aneurysmal subarachnoid hemorrhage have been seldom studied. On multivariable analysis, only presence of symptoms ascribed to vasospasm (PϽ0.01) and evidence of vasospasm on TCD or angiogram predicted cerebral infarction (PϽ0.01). TCD and angiogram agreed on the diagnosis of vasospasm in 73% of cases (95% CI, 63% to 81%), but the diagnostic accuracy of this combination of tests was suboptimal for the prediction of cerebral infarction occurrence (sensitivity, 0.72; specificity, 0.68; positive predictive value, 0.67; negative predictive value, 0.72). Location of the cerebral infarction on delayed CT was predicted by neurological symptoms in 74%, by aneurysm location in 77%, and by angiographic vasospasm in 67%. Conclusions-Evidence of vasospasm on TCD and angiogram is predictive of cerebral infarction on CT scan but sensitivity and specificity are suboptimal. Cerebral infarction location cannot be predicted in one quarter to one third of patients by any of the studied clinical or radiological variables. (Stroke.

A fundamental and shared process in all forms of life is the use of DNA glycosylase enzymes to excise rare damaged bases from genomic DNA. Without such enzymes, the highly-ordered primary sequences of genes would rapidly deteriorate. Recent structural and biophysical studies are beginning to reveal a fascinating multistep mechanism for damaged base detection that begins with short-range sliding of the glycosylase along the DNA chain in a distinct conformation we refer to as the search complex (SC). Sliding is frequently punctuated by the formation of a transient "interrogation" complex (IC) where the enzyme extrahelically inspects both normal and damaged bases in an exosite pocket that is distant from the active site. When normal bases are presented in the exosite, the IC rapidly collapses back to the SC, while a damaged base will efficiently partition forward into the active site to form the catalytically competent excision complex (EC). Here we review the unique problems associated with enzymatic detection of rare damaged DNA bases in the genome, and emphasize how each complex must have specific dynamic properties that are tuned to optimize the rate and efficiency of damage site location.The problem of enzymatic detection of a single damaged base in the context of a vast genome of nearly isomorphous undamaged bases has intrigued the DNA repair community virtually since the discovery of the DNA base excision repair pathway (1). The initiating step in this pathway begins with the enzymatic hydrolysis of the glycosidic bond that attaches the damaged base to the deoxyribose phosphate DNA backbone, setting the stage for the multistep base excision repair process to begin (Fig. 1). The cellular sentinels at this first step are the remarkable DNA glycosylase enzymes (2). Although these enzymes fall into different structural classes, and each is specialized for the detection and removal of different types of damaged bases (Table 1), with the sole exception of pyrimidine dimer DNA glycosylase (3), these enzymes have converged on a single mechanistic solution for damaged base recognition and excision: rotation of the damaged base from the DNA base stack into a sequestered active site pocket where chemistry occurs (Fig. 2). This process has been called either base or nucleotide "flipping" by various investigators (4,5), and connects the damage encounter event with the catalytic step of bond scission.Base flipping involves one of the most extended and improbable reaction trajectories in biology. The overall reaction is driven forward solely by the use of enzyme binding energy for DNA, which is used to pay for the significant energetic costs of extracting a base from the DNA base stack (11). These costs include breaking of Watson-Crick hydrogen bonds, the disruption of aromatic stacking interactions with adjacent bases, and large perturbations in the phosphate torsion angles around the flipped base. Initial structural investigations into † This work was supported by NIH grant GM056834 (J.T.S).

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Posttraumatic cerebrospinal fluid (CSF) leakage frequently complicates skull base fractures. While most CSF leaks will cease without treatment, patients with persistent CSF leaks may be at increased risk for meningitis, and many will require surgical intervention. We reviewed the medical records of 51 patients treated between 1984 and 1998, with CSF leaks that persisted for 24 hours or longer after head trauma. Twenty-eight patients (53%) had spontaneous resolution of the leakage at an average of 5 days. Twenty-three patients (47%) required surgery. Eight patients (16%) had occult leaks presenting with recurrent meningitis at an average of 6.5 years posttrauma. Forty-three (84%) patients with CSF leaks had an associated skull fracture, most commonly involving the frontal sinus, while only 18 patients (35%) had parenchymal brain injury or extra-axial hematoma. Eight patients (16%) had delayed leaks at an average of 13 days posttrauma. Among patients with clinically evident CSF leakage the frequency of meningitis was 10% with antibiotic prophylaxis, and 21% without antibiotic prophylaxis. Thus, prophylactic antibiotic administration halved risk of meningitis. A variety of surgical approaches was used, with minimal morbidity. Three of 23 surgically treated patients (13%) required additional surgery for continued leakage. Patients with CSF leaks that persist greater than 24 hours are at risk for meningitis, and many will require surgical intervention. Prophylactic antibiotics may be effective and should be considered in this group of patients. Patients with skull fractures involving the skull base or frontal sinus should be followed for delayed leakage. Surgical outcome is excellent.

RCH most commonly follows supratentorial neurosurgical procedures, performed with the patient in the supine position, that involve opening of cerebrospinal fluid cisterns or the ventricular system (such as unruptured aneurysm repair or temporal lobectomy). Preoperative aspirin use and moderately elevated intraoperative systolic blood pressure are potentially modifiable risk factors associated with the development of RCH. Although RCH can cause death or major morbidity, most cases are asymptomatic or exhibit a benign course. Cerebellar "sag" as a result of cerebrospinal fluid hypovolemia, causing transient occlusion of superior bridging veins within the posterior fossa and consequent hemorrhagic venous infarction, is the most likely pathophysiological cause of RCH.

Background and Purpose-Patients with hereditary hemorrhagic telangiectasia (HHT) are at risk for developing cerebral vascular malformations and pulmonary arteriovenous fistulae. We assessed the risk of neurological dysfunction from these malformations and fistulae. Methods-Three hundred twenty-one consecutive patients with HHT seen at a single institution over a 20-year period were studied. Any evidence of prior neurological symptoms or presence of an intracranial vascular malformation was recorded. All cases of possible cerebral arteriovenous malformation were confirmed by conventional arteriography. Results-Twelve patients (3.7%) had a history of cerebral vascular malformations. Ten patients had arteriovenous malformations, 1 had a dural arteriovenous fistula, and 1 had a cavernous malformation. Seven patients (2.1%) presented with intracranial hemorrhage, 2 presented with seizures alone, and 3 were discovered incidentally. The average age at the time of symptomatic intracranial hemorrhage was 25.4 years. All patients with a history of intracranial hemorrhage were classified as Rankin grade I or II at a mean follow-up interval of 6.0 years. A history of cerebral infarction or transient ischemic attack was found in 29.6% of patients with HHT and a pulmonary arteriovenous fistula. Conclusions-The risk of intracranial hemorrhage is low among people with HHT. Furthermore, a majority of these patients have a good functional outcome after hemorrhage. The data do not suggest a compelling indication for routine screening of patients with HHT for asymptomatic cerebral vascular malformations. 1 The estimated prevalence is 1 or 2 cases per 100 000. 1,2 Chromosomes 9q and 12q have been implicated in the inheritance of this disease. 3-7 A family history of HHT, although not required to make the diagnosis, is almost universally present. The vascular malformations of HHT may occur in multiple organs, including the lung, liver, kidney, and brain. 8,9 Epistaxis is the most common presentation, followed by gastrointestinal bleeding. 10,11 Pulmonary arteriovenous fistulae (AVFs) are often accompanied by dyspnea or hemoptysis and, less frequently, ischemic neurological symptoms. [12][13][14] Telangiectasias are frequently noted on the skin and mucous membranes, often not appearing until the second or third decade. 8,9 Brown et al 15,16 have reported that the detection rate of intracranial vascular malformations in a population-based study in the most recent time period was 2.75 per 100 000 person-years. The prevalence of intracranial hemorrhage from a vascular malformation was 7.50 per 100 000 people in the population at large. 15,16 Patients with HHT are thought to be at increased risk for harboring cerebral vascular malformations compared with the entire population. Arteriovenous malformations (AVMs), cavernous malformations, dural arteriovenous fistulae, and aneurysms have all been reported in these patients ( Figure 1). 14,17-24 Vascular malformations are found throughout the central nervous system, including the spinal cor...

We report an "exchange-rate-filtered" magnetic resonance approach that allows the detection of exchangeable protons of low concentration solutes without interference of non-exchanging protons. This indirect detection of signals of multiple rapidly exchanging protons through the water signal can be achieved while retaining chemical shift specificity and increasing sensitivity by several orders of magnitude with respect to standard spectroscopy. This frequency labeled exchange (FLEX) transfer principle is applied to detect previously "invisible" protons of some nucleic acids, and peptides, as well as rapidly exchanging protons (k > 300s −1 ) in so-called chemical exchange saturation transfer (CEST) MRI contrast agents. FLEX methodology is expected to provide a practical approach for the study of highly dynamic regions of nucleic acids and proteins where amide, amino and imino groups are rapidly moving between a closed solvent inaccessible state and an exposed state where exchange occurs. This alternative type of labeling and detecting exchangeable protons is also expected to greatly benefit the development of new exchange-based MRI contrast agents, providing a method for multi-frequency detection using frequency transfer instead of saturation transfer.Exchangeable protons have played an invaluable role in NMR studies of protein and nucleic acid structure and dynamics.1 -3 Detection of such solute protons (s) is governed by the "slow exchange" condition, where the frequency difference with water protons, Δω sw , is larger than the exchange rate, k s . However, the broad resonances of such protons are often "invisible", because they are either obscured by larger and narrower signals of other protons, or hidden in the noise. We report an "exchange-rate-filtered" approach that allows the detection of such pvanzijl@mri.jhu.edu. Supporting Information Available: Determination of labeling efficiency and details of data analysis procedures. This material is available free of charge via the Internet at http://pubs.acs.org. protons without interference of non-exchanging protons. Also, the transfer of these protons to water is exploited to achieve a sensitivity enhancement of several orders of magnitude with respect to standard spectroscopy. This new frequency labeled exchange (FLEX) transfer principle is applied to detect previously "invisible" protons of some nucleic acids and peptides, as well as rapidly exchanging protons (k s > 300s −1 ) in so-called chemical exchange saturation transfer (CEST) MRI contrast agents. 4-7 Using the FLEX approach, this latter detection can be achieved without the need for proton saturation and without need for additional postprocessing to separate the agent signals from background signals due to direct water saturation or due to interfering slower magnetization transfer effects. NIH Public AccessThe basic FLEX pulse sequence ( Fig. 1) consists of a series of label-transfer modules (LTMs) in which exchangeable solute protons are frequency labeled and subsequently transferred to water...