Mandibular prognathism attenuates brain blood flow induced by chewing.

Mastication is closely related to brain function. Animal experiments have revealed that tooth loss has a negative influence on brain function. Clinical studies also suggest that normal occlusion is an essential factor for favorable brain function. Mandibular prognathism (MP) usually results in occlusal dysfunction. However, the relationship between MP and brain function remains unclear. In the present study, we examined the relationship between MP and brain function by measuring brain blood flow (BBF). Seventeen subjects with normal occlusion (NORM) and 25 patients with MP participated in this study. The number of occlusal contacts were counted. Electromyography of the masseter muscles during clenching was also recorded. BBF was measured with non-invasive functional near-infrared spectroscopy during calculation task and chewing task. The number of the occlusal contacts and masseter muscle activity were lower in MP compared with NORM. The calculation task increased BBF in both groups. The chewing task also increased BBF in the inferior frontal gyrus in both groups, although the increase in MP was smaller than in NORM. We discovered that patients with MP exhibited a smaller increase in BBF at the inferior frontal gyrus during chewing as compared with NORM. As such, MP would negatively affect brain function.

Chemical synthesis of erythropoietin glycoforms for insights into the relationship between glycosylation pattern and bioactivity.

The role of sialyloligosaccharides on the surface of secreted glycoproteins is still unclear because of the difficulty in the preparation of sialylglycoproteins in a homogeneous form. We selected erythropoietin (EPO) as a target molecule and designed an efficient synthetic strategy for the chemical synthesis of a homogeneous form of five EPO glycoforms varying in glycosylation position and the number of human-type biantennary sialyloligosaccharides. A segment coupling strategy performed by native chemical ligation using six peptide segments including glycopeptides yielded homogeneous EPO glycopeptides, and folding experiments of these glycopeptides afforded the correctly folded EPO glycoforms. In an in vivo erythropoiesis assay in mice, all of the EPO glycoforms displayed biological activity, in particular the EPO bearing three sialyloligosaccharides, which exhibited the highest activity. Furthermore, we observed that the hydrophilicity and biological activity of the EPO glycoforms varied depending on the glycosylation pattern. This knowledge will pave the way for the development of homogeneous biologics by chemical synthesis.

Safe and efficient Boc-SPPS for the synthesis of glycopeptide-α-thioesters.

Boc-solid phase peptide synthesis is useful for the preparation of peptide-α-thioesters. However, this strategy usually employs hydrogen fluoride for the final deprotection step. These strongly acidic conditions cannot be applied for the synthesis of acid-labile glycopeptide-α-thioesters. The protocol presented here is a modified in situ neutralization Boc-solid phase peptide synthesis employing 10% sulfuric acid/dioxane conditions for intermediate Boc removal and TfOH for the final side-chain deprotection step. These conditions were found to be applicable for the synthesis of acid-labile glycopeptide-α-thioesters. In this protocol, a glycopeptide is synthesized as α- thioester on a thiol linker, and the product glycopeptide-α-thioester is released from the resin by thiolysis after side-chain deprotection step with an acid cocktail containing TfOH instead of hydrogen fluoride.

Mechanism of 4-nitrophenol oxidation in Rhodococcus sp. Strain PN1: characterization of the two-component 4-nitrophenol hydroxylase and regulation of its expression.

4-Nitrophenol (4-NP) is a toxic product of the hydrolysis of organophosphorus pesticides such as parathion in soil. Rhodococcus sp. strain PN1 degrades 4-NP via 4-nitrocatechol (4-NC) for use as the sole carbon, nitrogen, and energy source. A 5-kb EcoRI DNA fragment previously cloned from PN1 contained a gene cluster (nphRA1A2) involved in 4-NP oxidation. From sequence analysis, this gene cluster is expected to encode an AraC/XylS family regulatory protein (NphR) and a two-component 4-NP hydroxylase (NphA1 and NphA2). A transcriptional assay in a Rhodococcus strain revealed that the transcription of nphA1 is induced by only 4-NP (of several phenolic compounds tested) in the presence of nphR, which is constitutively expressed. Disruption of nphR abolished transcriptional activity, suggesting that nphR encodes a positive regulatory protein. The two proteins of the 4-NP hydroxylase, NphA1 and NphA2, were independently expressed in Escherichia coli and purified by ion-exchange chromatography or affinity chromatography. The purified NphA2 reduced flavin adenine dinucleotide (FAD) with the concomitant oxidation of NADH, while the purified NphA1 oxidized 4-NP into 4-NC almost quantitatively in the presence of FAD, NADH, and NphA2. This functional analysis, in addition to the sequence analysis, revealed that this enzyme system belongs to the two-component flavin-diffusible monooxygenase family. The 4-NP hydroxylase showed comparable oxidation activities for phenol and 4-chlorophenol to that for 4-NP and weaker activities for 3-NP and 4-NC.