Phase determination is one of the most difficult step for crystal structure determination. Many techniques, such as isomorphous replacement, multiple- or single-wavelength anomalous diffraction, molecular replacement, direct method, and their combinations, are often used for structure determination of biological macromolecules. Basic theories of phase determination of biological macromolecular crystallography are presented for students who want to learn a“Black Box”of phasing.
Pathogenic bacteria that produce metallo-β-lactamases (MBLs) are emerging as a new threat to the medical community. These enzymes catalyze the hydrolysis of almost β-lactam antibiotics. Moreover, serine-β-lactamase inhibitors are ineffective against MBLs. Thus, the development of inhibitors for MBLs is important. We prepared two inhibitors, pentafluorophenyl 3-mercaptopropionate (PFMP, 1) and 3- (3-mercaptopropionylsulfanyl) propionic acid pentafluorophenyl ester (MPAP, 2) for one of MBLs, IMP-1. From the gel-filtration experiment of the enzyme-inhibitor complex, these compounds inhibited IMP-1 irreversibly. Moreover, X-ray crystallography revealed that inhibitor 2 covalently binds to IMP-1 to form an amide bond between the amino group (Nζ) of Lys224 and the inhibitor.
The crystal structure of the P-protein of the glycine cleavage system from Thermus thermophilus HB8 has been determined, which reveals that P-proteins do not involve the α2 dimer universally observed in the evolutionarily related pyridoxal 5'-phosphate (PLP) -dependent enzymes. Instead, novel αβ dimers associate to form an α2β2 tetramer. The crystal structure of P-protein without PLP has also been analyzed. These structures allow the identification of functionally important residues involved in recognition of the cofactor and substrates, and show that most of these residues are well conserved among all living organisms. These results provide insights into the molecular basis of hyperglycinemia.
A novel crystal mounting technique was developed for the sulfur SAD (S-SAD) method using longer wavelength X-rays. With this novel technique, the cryo-buffer and cryoloop surrounding the protein crystal can be removed before data collection to eliminate their X-ray absorption. Structures of several novel proteins have been solved by S-SAD using this mounting technique in combination with in-house chromium Kα radiation (2.29 Å) . Our novel crystal mounting technique is superior to the standard cryoloop mounting method for the measurement of small anomalous differences, and yields good results in sulfur substructure solution and phasing by the SAD method. These results indicate that the S-SAD method with a chromium source becomes more practical for macromolecular structure determination using our crystal mounting technique.
This review article describes recent developments in the studies of hoop-shaped cyclic benzenoid,  cyclophenacene, derived from functionalization of  fullerene. The authors mainly discuss background, history, synthetic strategy, structure, and properties of such cyclic π-conjugates systems. Theoretical studies of structures and properties of finite-length armchair carbon nanotubes are also described.
Phytobilins, linear tetrapyrrole pigments utilized for light-harvesting and light-sensing in photosynthetic organisms, are synthesized from biliverdin by ferredoxin-dependent bilin reductases (FDBRs) . We have determined the crystal structures of substrate-free and substrate-bound forms of PcyA, one such FDBR. These structures revealed the first tertiary structures of an FDBR family member. Based on these structures, we discuss the binding mechanisms of ferredoxin and substrate to PcyA. Moreover, we propose the reaction mechanism how PcyA controls the two-step reductions of biliverdin. Homology modeling of other FDBRs using PcyA structure may provide structural bases for the reaction mechanisms of other FDBRs.
CEL-III is a Ca2+-dependent animal lectin from the sea cucumber, Cucumaria echinata, which shows binding specificity for galactose-containing carbohydrates and exhibits hemolytic and hemagglutinating activities. We have determined its crystal structure at 1.7 Å resolution by using single isomorphous replacement with anomalous scattering of lead. CEL-III consists of N-terminal two carbohydrate-binding domains and a C-terminal domain, which is essential for oligomerization and binding to a membrane. CEL-III is the first structure with two β-trefoil folds containing five Ca2+ ions. Here, we report the structural features of CEL-III and discuss the mechanisms of recognition of carbohydrates and hemolysis.
A small crystal structure database containing only basic inorganic substances was constructed, aiming mainly at educational use. Its design and principle of data checking are described, followed by short explanations on softwares attached.