Changes in pancreatic exocrine function, fat and fibrosis in diabetes mellitus: analysis using MR imaging.

OBJECTIVE: To evaluate the relationships between hemoglobin A1c (HbA1c) levels with exocrine pancreatic function using cine-dynamic magnetic resonance cholangiopancreatography (MRCP) and the pancreatic parenchyma using fat-suppressed T1 mapping and the proton density fat fraction (PDFF). METHODS: Patients who underwent 3T-MRI and HbA1c measurement were retrospectively recruited. MRI included cine-dynamic MRCP with a spatially selective inversion-recovery (SS-IR) pulse, fat-suppressed Look-Locker T1 mapping and multiecho 3D Dixon-based PDFF mapping. The pancreatic exocrine secretion grade on cine-dynamic MRCP, T1 values, and PDFF were analyzed in non-diabetic (n = 32), pre-diabetic (n = 44) and diabetic (n = 23) groups defined using HbA1c. RESULTS: PDFF was weakly correlation with HbA1c (ρ = 0.30, p = 0.002). No correlations were detected between HbA1c and secretion grade (ρ = - 0.16, p = 0.118) or pancreatic parenchymal T1 (ρ = 0.13, p = 0.19). The secretion grade was comparable between the three groups. The T1 value was higher in diabetic (T1 = 1006.2+/- 224.8 ms) than in non-diabetic (T1 = 896.2+/- 86.3 ms, p = 0.010) and pre-diabetic (T1 = 870.1+/- 91.7 ms, p < 0.010) patients. The PDFF was higher in diabetic (FF = 11.8+/- 8.7 %) than in non-diabetic (FF = 6.8+/- 4.2 %, p = 0.014) patients. CONCLUSION: Pancreatic exocrine function, T1, and FF showed no correlation with HbA1c. Pancreatic T1 and fat fraction is increased in patients with Type 2 diabetes mellitus. ADVANCES IN KNOWLEDGE: This study demonstrates unaffected exocrine function in pre-diabetes and diabetes and confirms that pancreatic parenchymal T1 and FF are increased in patients with diabetes.

Symptoms that are associated with decreased pancreatic enzyme flow: MRCP assessment.

BACKGROUND: Pancreatic exocrine insufficiency is one of the causes of malabsorption syndrome. In many cases of malabsorption syndrome, pancreatic exocrine insufficiency can be treated with pancreatic enzyme replacement therapy. Therefore, it is important to detect pancreatic endocrine insufficiency as early and accurately as possible. Recent studies have shown that cine-dynamic MR cholangiopancreatography (MRCP) may be useful to evaluate pancreatic exocrine function PURPOSE: To identify abdominal symptoms that suggest decreased flow of pancreatic enzyme secretion for which cine-dynamic MRCP should be performed to diagnose pancreatic exocrine insufficiency. STUDY TYPE: Prospective. POPULATION: In all, 111 patients with various types of abdominal symptoms. FIELD STRENGTH/SEQUENCE: 5 T or 3 T, MRCP with spatially selective inversion recovery pulse (cine-dynamic MRCP). ASSESSMENT: Cine-dynamic MRCP was performed and an 18-question clinical questionnaire on abdominal symptoms was administered. The secretion grade derived from cine-dynamic MRCP was compared between those answering "yes" and "no" for all 18 items STATISTICAL TESTS: Univariate analysis and further analyzed using multiple regression analysis. The associations between the secretion grade and the items in the clinical questionnaire were analyzed by univariate analysis and further analyzed using multiple regression analysis. RESULTS: The following three items showed significantly negative correlations with secretion grade: Q9, Does your rectal gas smell foul? (ß = -0.44, P = 0.001); Q13, Is stool quantity large? (ß = -0.41, P = 0.001); and Q18, Are your stools soft? (ß = -0.53, P < 0.001). No significant correlations with exocrine pancreatic function measured by cine-dynamic MRCP were seen for the remaining 15 abdominal symptom items. DATA CONCLUSION: Abdominal symptoms that suggest decreased flow of pancreatic enzyme secretion were foul rectal gas, large stool, and soft stool. Pancreatic exocrine insufficiency due to decreased pancreatic enzyme flow may be suspected in patients with these abdominal symptoms. LEVEL OF EVIDENCE: 1 Technical Efficacy Stage: 3 J. Magn. Reson. Imaging 2019;50:417-423.

Alterations in apparent diffusion coefficient values of the kidney during the cardiac cycle: Evaluation with ECG-triggered diffusion-weighted MR imaging.

PURPOSE: To evaluate dynamic changes in apparent diffusion coefficient (ADC) values of the kidney at different time points during the cardiac cycle using electrocardiographic (ECG)-triggered diffusion-weighted MR imaging in normal subjects, and to elucidate the differences in ADC values between the right and left kidneys during a cardiac cycle. MATERIALS AND METHODS: The study was approved by our institutional review board and informed consent was obtained from subjects. Twenty healthy volunteers who underwent ECG-triggered diffusion-weighted MR imaging of the kidney were included. The differences in ADC values of each kidney during different cardiac phases were compared. Additionally, the differences in maximum and minimum ADC values between the right and left kidney were also evaluated. RESULTS: ADC values in the right and left kidney changed significantly during the cardiac cycle (P < 0.00001). Maximum and minimum ADC values during the cardiac cycle of the left kidney were significantly higher (P = 0.026 and 0.017, respectively) than those of the right kidney. Maximum ADC value in the left kidney had a significantly strong positive correlation with the left renal vein ratio (r = 0.83, P < 0.00001). In the right kidney, maximum ADC showed a weakly positive correlation with the diameter of the right renal vein (r = 0.45, P = 0.048). CONCLUSION: ADC values of the kidney obtained using ECG-triggered diffusion-weighted MR imaging change significantly during the cardiac cycle. Maximum (systolic) ADC during the cardiac cycle of the left kidney was significantly higher than that of the right kidney, probably due to the anatomical difference in the renal vein.

Postprandial changes in secretory flow of pancreatic juice in the main pancreatic duct: evaluation with cine-dynamic MRCP with a spatially selective inversion-recovery (IR) pulse.

PURPOSE: To evaluate the influence of oral ingestion on the secretory flow dynamics of physiological pancreatic juice within the main pancreatic duct in healthy subjects by using cine-dynamic MRCP with spatially-selective inversion-recovery (IR) pulse non-invasively. MATERIALS AND METHODS: Thirty-eight healthy subjects were investigated. MRCP with spatially-selective IR pulse was repeated every 15 s for 5 min to acquire a total of 20 images (cine-dynamic MRCP). A set of 20 MRCP images was repeatedly obtained before and after liquid oral ingestion every 7 min (including 2-min interval) for 40 min (a total of seven sets). Secretion grade of pancreatic juice on cine-dynamic MRCP was compared before and after oral ingestion using the nonparametric Wilcoxon signed-rank test. RESULTS: Median secretion grades of pancreatic juice at 5 min (score = 2.15), 12 min (score = 1.95) and 19 min (score = 2.05) after ingestion were significantly higher than that before ingestion (score = 1.40) (P = 0.004, P = 0.032, P = 0.045, respectively). Secretion grade of pancreatic juice showed a maximum peak of 2.15 at 5 min after ingestion. Thereafter, the secretion grade of pancreatic juice tended to gradually decline. CONCLUSION: Non-invasive cine-dynamic MRCP using spatially-selective IR pulse showed potential for evaluating postprandial changes in the secretory flow dynamics of pancreatic juice as a physiological reaction. KEY POINTS: • Secretion grade of pancreatic juice at cine-dynamic MRCP after ingestion was evaluated. • Secretion grade was significantly increased within 19 min after liquid meal ingestion. • Secretion grade showed maximum peak of 2.15 at 5 min after ingestion. • Postprandial changes in pancreatic juice flow can be assessed by cine-dynamic MRCP.

Pancreatic adenocarcinomas without secondary signs on multiphasic multidetector CT: association with clinical and histopathologic features.

PURPOSE: To determine the clinical, histopathologic and imaging features of pancreatic adenocarcinomas without secondary signs on dynamic CT. MATERIALS AND METHODS: Seventy patients (mean age 70 years) with histologically proven pancreatic adenocarcinoma underwent preoperative contrast material-enhanced multiphasic multidetector CT before pancreatic resection. In each patient, clinical data including carbohydrate antigen 19-9, frequency of isoattenuating tumours, and presence of secondary signs and histopathologic findings such as tumour location, tumour stage, and microscopic infiltrative growth grade were evaluated. RESULTS: Ten tumours (14%) were without secondary signs, and 60 (86%) were with secondary signs. Tumours without and with secondary signs were located in the uncinate process in 5 (50%) and 3 (5%), head in 3 (30%) and 29 (48%), body in 2 (20%) and 22 (37%), and tail in 0 (0%) and 6 (10%), respectively (p = .001). The frequency of isoattenuating pancreatic adenocarcinomas without secondary signs was significantly higher than those with secondary signs (p = 0.034). The tumour stage of pancreatic adenocarcinomas without secondary signs was earlier than that in tumours with secondary signs (p = 0.041). CONCLUSIONS: Pancreatic adenocarcinomas without secondary signs is characterized by the presence of uncinate and isoattenuating tumours and earlier tumour stage compared to tumours with secondary signs. KEY POINTS: Frequency of pancreatic adenocarcinomas without secondary signs on multiphasic CT is 14 . Pancreatic adenocarcinomas without secondary signs are common in the uncinate process. Pancreatic adenocarcinomas without secondary signs are common in isoattenuating tumours. Pancreatic adenocarcinomas without secondary signs are characterized by earlier-stage tumours.

Noninvasive investigation of exocrine pancreatic function: Feasibility of cine dynamic MRCP with a spatially selective inversion-recovery pulse.

PURPOSE: To investigate the feasibility of noncontrast-enhanced cine dynamic magnetic resonance cholangiopancreatography (MRCP) with a spatially selective inversion-recovery (IR) pulse for evaluating exocrine pancreatic function in comparison with the N-benzoyl-L-tyrosyl-p-aminobenzoic acid (BT-PABA) test as a pancreatic exocrine function test. MATERIALS AND METHODS: Twenty subjects with or without chronic pancreatitis were included. MRCP with a spatially selective IR pulse was repeated every 15 seconds for 5 minutes to acquire a total of 20 images (cine-dynamic MRCP). The median and mean frequency of the observation (the number of times) and the moving distance (mean secretion grading scores) of pancreatic juice inflow on cine-dynamic MRCP were compared with a BT-PABA test. RESULTS: The urinary PABA excretion rate (%) had significant positive correlations with both the mean secretion grade (r = 0.66, P = 0.002) and frequency of secretory inflow (r = 0.62, P = 0.004) in cine dynamic MRCP. Both the mean frequency of observations of pancreatic secretory inflow (1.4 ± 1.6 times vs. 14.3 ± 4.2 times, P < 0.001) and the mean secretion grade (grade = 0.16 ± 0.24 vs. grade = 1.81 ± 0.81, P < 0.001) was significantly lower in the chronic pancreatitis group than in the normal subject group. CONCLUSION: Cine dynamic MRCP with a spatially selective IR pulse may have potential for estimating the pancreatic exocrine function noninvasively as a substitute for the BT-PABA test.

Magnetisation transfer MR imaging of the kidney: evaluation at 3.0 T in association with renal function.

OBJECTIVE: To evaluate the feasibility of using magnetisation transfer (MT) MRI of the kidney at 3.0 T to assess renal function. METHODS: Forty-four patients who underwent abdominal MRI on a 3.0-T system including gradient-echo (GRE) sequences with and without MT pulse were included. In each patient, MT ratio (MTR) of the renal cortex and medulla was measured by using regions of interest (ROIs) placed on the MTR map image. RESULTS: Regression analysis showed good correlation between estimated glomerular filtration rate (eGFR) and MTR of the renal cortex (r = -0.645, P < 0.0001). Among 44 patients, 22 were categorised as the normal renal function group and 22 were classified as the decreased eGFR group. The mean MTR of the renal cortex in patients with decreased eGFR (mean MTR, 30.7 ± 3.2 %) was significantly higher (P < 0.0001) than that in patients with normal renal function (mean MTR, 25.3 ± 2.2 %), although the mean MTRs of the renal medulla in the two groups were not significantly different. CONCLUSION: There was good correlation between eGFR and MTR of the renal cortex derived from MT MRI at 3.0 T. This technique may have the potential to evaluate the degree of renal function non-invasively in patients with renal impairment. KEY POINTS: • Magnetisation transfer techniques can provide new information about renal disease. • MTR values of the renal cortex correlate well with estimated glomerular filtration. • Higher MTR of the renal cortex exists in patients with renal dysfunction. • MT MRI at 3.0 T may be useful for evaluating renal function.

Small hepatocellular carcinomas in cirrhosis: differences in contrast enhancement effects between helical CT and MR imaging during multiphasic dynamic imaging.

PURPOSE: The purpose of this study was to evaluate differences in the degrees of contrast enhancement effects of small hepatocellular carcinomas (HCCs) in patients with cirrhosis between helical computed tomography (CT) and magnetic resonance (MR) imaging during multiphasic contrast-enhanced dynamic imaging and to determine the diagnostic value of MR imaging especially in assessing hypovascular HCCs detected as hypoattenuating nodules on late-phase CT. SUBJECTS AND METHODS: This study included 64 small HCCs (<3 cm in diameter) in 40 patients with chronic hepatitis or cirrhosis who underwent multiphasic (arterial, portal and late phases) contrast-enhanced dynamic helical CT and MR imaging. The contrast enhancement patterns of each lesion in the arterial and late phases were evaluated by two radiologists experienced in liver MR imaging and categorized as one of five grades (1=hypoattenuated/hypointense; 2=slightly hypoattenuated/hypointense; 3=isoattenuated/isointense; 4=slightly hyperattenuated/hyperintense; 5=hyperattenuated/hyperintense), compared with the surrounding liver parenchyma. RESULT: Forty-three (67%) of 64 lesions showed Grade 4 (n=24) or Grade 5 (n=19) enhancement on arterial-phase CT, while 51 (80%) of 64 lesions showed Grade 4 (n=20) or Grade 5 (n=31) enhancement on arterial-phase MR imaging, indicating hypervascular HCCs. The grading score of hypervascular HCCs on arterial-phase MR imaging (mean: 4.61) was significantly (P<.01) higher than that for hypervascular HCCs on arterial-phase CT (mean: 4.20), showing better detection of the hypervascularity (arterial enhancement) of the lesion on arterial-phase MR imaging. Regarding hypovascular HCCs, all (100%) of 21 hypovascular HCCs on CT showed Grade 1 (n=10) or Grade 2 (n=11) enhancement on late-phase CT, seen as hypoattenuation. In contrast, 8 (62%) of 13 hypovascular HCCs on MR imaging showed Grade 1 (n=1) or Grade 2 (n=7) enhancement on late-phase MR imaging, seen as hypointensity. Grading scores of hypovascular HCCs on late-phase images were significantly (P<.001) lower on CT than on MR imaging (mean score: 1.52 vs. 2.31), indicating better washout effects for hypovascular HCCs on late-phase CT. CONCLUSION: The washout effects for small HCCs on late-phase MR imaging were inferior to those for small HCCs on late-phase CT. Especially, hypovascular HCCs demonstrated as hypoattenuating nodules on late-phase CT were often not seen on late-phase MR imaging, requiring careful evaluation of other sequences, including unenhanced T(1)-weighted and T(2)-weighted MR images.

Small hyperintense hepatic lesions on T1-weighted images in patients with cirrhosis: evaluation with serial MRI and imaging features for clinical benignity.

PURPOSE: The aim of this study was to evaluate the frequency and magnetic resonance imaging (MRI) features of clinically benign, small (<2 cm) hyperintense hepatic lesions in the cirrhotic liver on T1-weighted MR images seen at serial MRI. MATERIALS AND METHODS: This study included 189 patients with cirrhosis, who underwent hepatic MRI more than twice with an interval of at least 12 months. The initial MR images were reviewed for the presence of small hyperintense lesions on T1-weighted images. The size, location and signal intensity on T2-weighted images as well as enhancement patterns of the corresponding lesions were recorded. RESULTS: On the initial T1-weighted MR images, 43 small hyperintense hepatic lesions were detected in 23 (12%) of 189 patients. Twelve (28%) of 43 lesions showed early enhancement and were pathologically diagnosed as hepatocellular carcinoma (HCC) during the follow-up period. Thirty-one (72%) of 43 lesions showed no early enhancement with various signal intensity on T2-weighted images (hyperintensity=4, isointensity=20, hypointensity=7). Among these 31 lesions, 12 showed no interval change, while 11 disappeared (n=10) or decreased in size (n=1). In the remaining eight lesions, seven were diagnosed as HCC on the basis of pathologic confirmation or the interval growth. CONCLUSION: Small hyperintense hepatic lesions on T1-weighted magnetic resonance (MR) images without early enhancement on the arterial-phase contrast-enhanced dynamic studies in patients with cirrhosis usually showed no interval growth or disappeared during the serial MRI. These lesions with additional findings of iso- or hypointensity on the T2-weighted MR images without "washout effect" on the contrast-enhanced equilibrium-phase images may more frequently be clinically benign or hyperplastic nodules than HCCs.

Small hepatic lesions found on single-phase helical CT in patients with malignancy: diagnostic capability of breath-hold, multisection fluid-attenuated inversion-recovery (FLAIR) MR imaging using a half-fourier acquisition single-shot turbo spin-echo (HASTE) sequence.

PURPOSE: To evaluate the diagnostic capability of breath-hold, multisection fluid-attenuated inversion-recovery (FLAIR) imaging using a half-Fourier acquisition single-shot turbo spin-echo (HASTE) sequence in combination with T2-weighted fast spin-echo (FSE) magnetic resonance (MR) sequences for small hepatic lesions found on CT in patients with malignancy. MATERIALS AND METHODS: This study included 48 patients with extrahepatic malignancy who underwent both CT and MR examinations. There were a total of 112 small hepatic lesions (73 cysts and 39 liver metastases, <2 cm in diameter) that showed low attenuation on enhanced CT. Three radiologists independently reviewed the CT and MR (FLAIR-HASTE and T2-weighted FSE) images and assigned a confidence level to their evaluation (cyst or metastasis) on a five-point scale. RESULTS: All three reviewers were significantly better able (P < 0.05) to differentiate small hepatic cyst from liver metastasis with combined FLAIR-HASTE and T2-weighted FSE images (Az values = 0.997-0.999) than with CT (0.917-0.932). The mean values of sensitivity, specificity, and accuracy were significantly higher (P < 0.001) for T2-weighted FSE with FLAIR-HASTE (96.6%, 96.8%, and 96.7%, respectively) than for CT (76.9%, 61.6%, and 67.3%, respectively). A confident diagnosis was rendered in 12 of 112 lesions (10.7%) on the basis of CT, and this rate increased to 83 of 112 (74.1%) on the basis of T2-weighted FSE and FLAIR-HASTE imaging. CONCLUSION: FLAIR-HASTE is considered to be an effective sequence for differentiating hepatic cysts from liver metastases without the use of a contrast agent. With FLAIR-HASTE one can confidently diagnose small hepatic lesions found on CT in patients with a malignancy.

Structure of the carboxypeptidase Y inhibitor IC in complex with the cognate proteinase reveals a novel mode of the proteinase-protein inhibitor interaction.

Carboxypeptidase Y (CPY) inhibitor, IC, shows no homology to any other known proteinase inhibitors and rather belongs to the phosphatidylethanolamine-binding protein (PEBP) family. We report here on the crystal structure of the IC-CPY complex at 2.7 A resolution. The structure of IC in the complex with CPY consists of one major beta-type domain and a N-terminal helical segment. The structure of the complex contains two binding sites of IC toward CPY, the N-terminal inhibitory reactive site (the primary CPY-binding site) and the secondary CPY-binding site, which interact with the S1 substrate-binding site of CPY and the hydrophobic surface flanked by the active site of the enzyme, respectively. It was also revealed that IC had the ligand-binding site, which is conserved among PEBPs and the putative binding site of the polar head group of phospholipid. The complex structure and analyses of IC mutants for inhibitory activity and the binding to CPY demonstrate that the N-terminal inhibitory reactive site is essential both for inhibitory function and the complex formation with CPY and that the binding of IC to CPY constitutes a novel mode of the proteinase-protein inhibitor interaction. The unique binding mode of IC toward the cognate proteinase provides insights into the inhibitory mechanism of PEBPs toward serine proteinases and into the specific biological functions of IC belonging to the PEBP family as well.

Crystallization and preliminary X-ray analysis of carboxypeptidase Y inhibitor IC complexed with the cognate proteinase.

Carboxypeptidase Y (CPY) inhibitor I(C) is a naturally occurring serine carboxypeptidase inhibitor from Saccharomyces cerevisiae, the sequence of which is not homologous with any other known proteinase inhibitor and is classified as the phosphatidylethanolamine-binding protein (PEBP). I(C) has been crystallized in complex with the deglycosylated form of CPY by the hanging-drop vapour-diffusion technique with ammonium sulfate as a precipitant. The crystals of the complex belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 81.13, b = 186.6, c = 65.14 A. Diffraction data were collected to 2.7 A resolution. Structure determination of the complex is in progress by the molecular-replacement method using the structure of CPY as a search model.

Structures of two lectins from the roots of pokeweed (Phytolacca americana).

Pokeweed lectin (PL), a lectin specific for N-acetylglucosamine-containing saccharides, stimulates peripheral lymphocytes to undergo mitosis by binding to their cell surfaces. Four types of lectins have been isolated from the roots of pokeweed (Phytolacca americana) and shown to contain homologous domains but to have different molecular sizes and biological properties. PL-D, the smallest lectin in the group, has two isolectins, PL-D1 and PL-D2. PL-D1 consists of 84 amino-acid residues, while PL-D2 is identical to PL-D1 in sequence except for the lack of two C-terminal residues, Leu83 and Thr84. The crystal structures of PL-D1 and PL-D2 were solved by the molecular-replacement method and refined to 1.65 and 1.5 A resolution with R factors of 17.2 and 17.6%, respectively. The PL-Ds are composed of two repetitive chitin-binding domains, each of which has four S-S bridges and one putative carbohydrate-binding site. The two carbohydrate-binding sites in PL-D are located on one side of the molecule. The relative orientation of the two domains in PL-D1 differs from that in PL-D2. Two C-terminal residues of PL-D1 are invisible in the present crystal structure, indicating the flexibility of the region. PL-D2 has a Ca2+ ion bound to the C-terminus on the molecular surface. A wide distribution of acidic residues is characteristically observed on one side of the C-terminal region of PL-D.

Similarity between protein-protein and protein-carbohydrate interactions, revealed by two crystal structures of lectins from the roots of pokeweed.

The roots of pokeweed (Phytolacca americana) are known to contain the lectins designated PL-A, PL-B, PL-C, PL-D1, and PL-D2. Of these lectins, the crystal structures of two PLs, the ligand-free PL-C and the complex of PL-D2 with tri-N-acetylchitotriose, have been determined at 1.8A resolution. The polypeptide chains of PL-C and PL-D2 form three and two repetitive chitin-binding domains, respectively. In the crystal structure of the PL-D2 complex, one trisaccharide molecule is shared mainly between two neighboring molecules related to each other by a crystallographic 2(1)-screw axis, and infinite helical chains of complexed molecules are generated by the sharing of ligand molecules. The crystal structure of PL-C reveals that the molecule is a dimer of two identical subunits, whose polypeptide chains are located in a head-to-tail fashion by a molecular 2-fold axis. Three putative carbohydrate-binding sites in each subunit are located in the dimer interface. The dimerization of PL-C is performed through the hydrophobic interactions between the carbohydrate-binding sites of the opposite domains in the dimer, leading to a distinct dimerization mode from that of wheat-germ agglutinin. Three aromatic residues in each carbohydrate-binding site of PL-C are involved in the dimerization. These residues correspond to the residues that interact mainly with the trisaccharide in the PL-D2 complex and appear to mimic the saccharide residues in the complex. Consequently, the present structure of the PL-C dimer has no room for accommodating carbohydrate. The quaternary structure of PL-C formed through these putative carbohydrate-binding residues may lead to the lack of hemagglutinating activity.

Crystallization and preliminary X-ray analysis of lectin C from the roots of pokeweed (Phytolacca americana).

Lectin C from the roots of pokeweed (Phytolacca americana) (PL-C; 13 747 Da, 126 amino-acid residues), which consists of three chitin-binding domains, was initially crystallized in two crystal forms. One form, obtained in the presence of 30%(w/v) PEG 4000, belongs to the tetragonal system. The other, obtained in the presence of 2.0 M ammonium sulfate, belongs to the rhombohedral system. Statistical analyses of the X-ray diffraction intensities showed that both crystals were twinned. Single crystals suitable for a diffraction experiment were obtained by the addition of 0.5%(v/v) dioxane to the latter precipitant solution. They belong to space group R3, with unit-cell parameters a = b = 104.1, c = 69.7 A, and diffract X-rays to 1.8 A resolution. A heavy-atom derivative crystal has been obtained and structure determination is presently ongoing using the SIRAS method.

The 1.6-A crystal structure of the copper(II)-bound bleomycin complexed with the bleomycin-binding protein from bleomycin-producing Streptomyces verticillus.

Bleomycin (Bm) in the culture broth of Streptomyces verticillus is complexed with Cu(2+) (Cu(II)). In the present study, we determined the x-ray crystal structures of the Cu(II)-bound and the metal-free types of Bm at a high resolution of 1.6 and 1.8 A, respectively, which are complexed with a Bm resistance determinant from Bm-producing S. verticillus, designated BLMA. In the current model of Cu(II).Bm complexed with BLMA, two Cu(II).Bm molecules bind to the BLMA dimer. The electron density map shows that the copper ion is clearly defined in the metal-binding domain of the Bm molecule. The metal ion is penta-coordinated by a tetragonal monopyramidal cage of nitrogens and binds to the primary amine of the beta-aminoalanine moiety of Bm. The binding experiment between Bm and BLMA showed that each of the two Bm-binding pockets has a different dissociation constant (K(d)(1) and K(d)(2)). The K(d)(1) value of 630 nm for the first Bm binding is larger than the K(d)(2) value of 120 nm, indicating that the first Bm binding gives rise to a cooperative binding of the second Bm to the other pocket.