Production and synthesis of a novel Pt-labeled platinum complex and evaluation of its biodistribution in healthy mice

Omokawa, Marina*; Kimura, Hiroyuki*; Hatsukawa, Yuichi*; Kawashima, Hidekazu*; Tsukada, Kazuaki; Yagi, Yusuke*; Naito, Yuki*; Yasui, Hiroyuki*

Bioorganic & Medicinal Chemistry, 97, p.117557_1 - 117557_6, 2024/01

https://doi.org/10.1016/j.bmc.2023.117557

Structure of HIV-1 protease in complex with potent inhibitor KNI-272 determined by high-resolution X-ray and neutron crystallography

Adachi, Motoyasu; Ohara, Takashi; Kurihara, Kazuo; Tamada, Taro; Honjo, Eijiro; Okazaki, Nobuo; Arai, Shigeki; Shoyama, Yoshinari; Kimura, Kaname*; Matsumura, Hiroyoshi*; et al.

Proceedings of the National Academy of Sciences of the United States of America, 106(12), p.4641 - 4646, 2009/03

https://doi.org/10.1073/pnas.0809400106

To further understand the catalytic mechanism and inhibitor recognition of HIV-1 protease, we need to determine the locations of key hydrogen atoms in the catalytic aspartates Asp25 and Asp125. The structure of HIV-1 protease in complex with transition-state analog KNI-272 was determined by combined neutron crystallography at 1.9 resolution and X-ray crystallography at 1.4 resolution. The resulting structural data shows that the catalytic residue Asp25 is protonated and that Asp125 is deprotonated. The proton on Asp25 makes a hydrogen bond with the carbonyl group of the allophenylnorstatine group in KNI-272. The deprotonated Asp125 bonds to the hydroxyl proton of Apns. The results provide direct experimental evidence for proposed aspects of the catalytic mechanism of HIV-1 protease; and can therefore contribute substantially to the development of specific inhibitors for therapeutic application.

Structures of drug-target proteins determined by both X-ray and neutron diffraction

Kuroki, Ryota

no journal, , 

X-ray and neutron crystallography enables us to obtain accurate atomic positions within proteins. The structure of porcine pancreatic elastase (PPE) with its potent inhibitor (FR130180) was determined to 0.94 resolution by X-ray diffraction and 1.75 resolution by neutron diffraction. It was found that there are two characteristic hydrogen bonding interactions in which hydrogen atoms were confirmed. One is located between a catalytic aspartate and histidine, another is involved in the inhibitor recognition site. The structure of HIV-PR with its potent inhibitor (KNI-272) was also determined to 0.93 resolution by X-ray diffraction and 2.3 resolution by neutron diffraction. The ionization state of the catalytic residues were clarified to show that Asp125 is protonated and Asp25 is deprotonated. The ionization state and the location of hydrogen atoms of the catalytic residue in HIV-PR were firstly determined by neutron diffraction.

Neutron crystal structure analysis of HIV-1 protease complexed with KNI-272

Adachi, Motoyasu; Ohara, Takashi; Kurihara, Kazuo; Tamada, Taro; Honjo, Eijiro; Okazaki, Nobuo; Arai, Shigeki; Shoyama, Yoshinari; Matsumura, Hiroyoshi*; Sugiyama, Shigeru*; et al.

no journal, , 

We have determined a crystal structure of HIV-1 protease by neutron crystallography. The development of HIV-1 protease inhibitors is regarded as a major success of structure-based drug design and contributes to establish highly active anti-retroviral therapy for AIDS. To further understand the catalytic mechanism of HIV-1 protease and interaction between HIV-1 protease and its inhibitor, we have determined the crystal structure of HIV-1 protease in complex with a inhibitor, KNI-272 to 2.3 resolution by neutron crystallography. Our results indicates that the carbonyl group of allophenylnorstatine (Apns) in KNI-272 forms a significant hydrogen bond with protonated Asp 25, and the hydrogen atom from the hydroxyl group of Apns forms a remarkable hydrogen bond with the deprotonated Asp125. These results show direct evidence that Asp25 provides a proton to carbonyl group of substrate and Asp125 contributes to activate the attacking water molecule as a nucleophile.

Reactivity of a spin trapping agent, CYPMPO, toward water decomposition radicals

Midorikawa, Masamichi*; Yamashita, Shinichi; Katsumura, Yosuke; Lin, M.; Muroya, Yusa*; Maeyama, Takuya*; Funtowiez, D.*; Kamibayashi, Masato*; Anzai, Kazunori*

no journal, , 

no abstracts in English

Measurement of the reactivity of a novel spin-trapping agent, CYPMPO, by pulse radiolysis study

Midorikawa, Masamichi*; Oka, Toshitaka; Yamashita, Shinichi; Muroya, Yusa*; Lin, M.; Kamibayashi, Masato*; Anzai, Kazunori*; Kudo, Hisaaki*; Katsumura, Yosuke

no journal, , 

5-(2,2-dimethyl-1,3-propoxy cyclophosphoryl)-5-methyl-1pyrroline N-oxide (CYPMPO) is a recently developed spin-trapping agent, which is expected to distinguish O from OH. We have determined its reactivity with water decomposed radicals, especially with OH and with e by pulse radiolysis, and compared results with commonly-used spin trapping agent, 5,5-dimethyl-1-pyrroline N-oxide (DMPO). Furthermore, the reactions with oxidative radicals such as COO, NO and Br have also been investigated.

X-ray structure analysis of A17 type HIV-1 protease complexed with inhibitors

Adachi, Motoyasu; Arai, Shigeki; Tamada, Taro; Kuroki, Ryota; Hidaka, Koshi*; Kimura, Toru*; Kiso, Yoshiaki*

no journal, , 

To obtain information for design of inhibitor against drug resistant mutant HIV protease, we determined X-ray crystal structures of A17 type HIV-1 protease complexed with inhibitors of lopinavir and KNI-1657. The gene of A17 type HIV-1 protease was synthesized chemically, and prepared using E. coli expression system. The results showed that affinity of lopinavir to the protease was 700 times lower than that of wild-type, and KNI-1657 has 20 times higher affinity to the protease than lopinavir. The crystals of complex were obtained using PEG4000 as precipitant. The diffraction data were collected at PF and SPring-8, and the structures were refined at R-factor of 19%.

Crystal structure analysis of HIV-1 protease by complementary use of synchrotron radiation and neutron

Adachi, Motoyasu; Ohara, Takashi; Kurihara, Kazuo; Tamada, Taro; Honjo, Eijiro*; Okazaki, Nobuo; Arai, Shigeki; Shoyama, Yoshinari*; Matsumura, Hiroyoshi*; Sugiyama, Shigeru*; et al.

no journal, , 

In this study, we determined crystal structures of HIV-1 protease complexed with inhibitor by neutron and X-ray crystallography. Finally, we refined the structures to R-factor of 17.3% and free R-factor 20.3% by neutron crystallography and to R-factor of 10.4 % and free R-factor 12.4% by X-ray crystallography. The result shows that Asp 25 residue is protonated and Asp 125 is deprotonated. These information is important to resolve catalytic mechanism and design of new potent inhibitor.

X-ray structure analysis of single-chained HIV-1 protease complexed with inhibitor

Adachi, Motoyasu; Tamada, Taro; Kuroki, Ryota; Hidaka, Koshi*; Kimura, Toru*; Kiso, Yoshiaki*; Yamamoto, Kohei*; Kidokoro, Shunichi*

no journal, , 

no abstracts in English

Use of substituted oxamide structure in designing pseudo-symmetric HIV protease inhibitors to employ multiple bridging water molecules

Hidaka, Koshi*; Toda, Yuki*; Adachi, Motoyasu; Kuroki, Ryota; Kiso, Yoshiaki*

no journal, , 

Molecular dynamic simulations of the inhibitor suggested existence of additional stable bridging water molecules to support the binding with mutated proteases. To increase the numbers of bridging water molecules, we replaced amide with sulfonyl and oxamide structures at both terminals of pseudo-symmetric peptides based on HMC. In summary, oxamide modifications resulted in a moderate activity loss against lopinavir-resistant mutations compared to inhibitors without oxamide. This method to accompany multiple bridging water molecules could be applicable to design protease inhibitors to defeat the drug resistance from amino acid mutations.