<scp>R</scp>aman Spectroscopy of Proteins

Kitagawa, Teizo; Hirota, Shun

doi:10.1002/0470027320.s8202

Cited by 41 publications

(63 citation statements)

References 217 publications

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“…1c, curve a), Raman bands at 662, 825, 1,017, 1,397, and 1,572 cm Ϫ1 are due to CaDPA (9,13), the band at 1,004 cm Ϫ1 is due to phenylalanine in spore protein, and the bands at 1,650 and 1,665 cm Ϫ1 are associated with ␣-helical and irregular structures of the protein amide I (peptide bond C ϭ O stretch), respectively (10). In examining the Raman spectra of spores that were wet heat treated at 95°C (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Thus, a detailed understanding of spore germination requires analyses of individual spores' germination behavior. Raman spectroscopy is widely used in biochemical studies, as this technique has high sensitivity and responds rapidly to subtle changes in molecular structure (10). When Raman spectroscopy is combined with confocal microscopy and optical tweezers, the resultant laser tweezers Raman spectroscopy (LTRS) allows the nondestructive, noninvasive detection of biochemical processes at the single-cell level (3,9,31).…”

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confidence: 99%

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Kinetics of Germination of Wet-Heat-Treated Individual Spores of Bacillus Species, Monitored by Raman Spectroscopy and Differential Interference Contrast Microscopy

Wang

Zhang

Setlow

et al. 2011

Appl Environ Microbiol

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Raman spectroscopy and differential interference contrast (DIC) microscopy were used to monitor the kinetics of nutrient and nonnutrient germination of multiple individual untreated and wet-heat-treated spores of Bacillus cereus and Bacillus megaterium, as well as of several isogenic Bacillus subtilis strains. Major conclusions from this work were as follows. (i) More than 90% of these spores were nonculturable but retained their 1:1 chelate of Ca 2؉ and dipicolinic acid (CaDPA) when incubated in water at 80 to 95°C for 5 to 30 min. (ii) Wet-heat treatment significantly increased the time, T lag , at which spores began release of the great majority of their CaDPA during the germination of B. subtilis spores with different nutrient germinants and also increased the variability of T lag values. (iii) The time period, ⌬T release , between T lag and the time, T release , at which a spore germinating with nutrients completed the release of the great majority of its CaDPA, was also increased in wet-heat-treated spores. (iv) Wet-heat-treated spores germinating with nutrients had higher values of I release , the intensity of a spore's DIC image at T release , than did untreated spores and had much longer time periods, ⌬T lys , for the reduction in I release intensities to the basal value due to hydrolysis of the spore's peptidoglycan cortex, probably due at least in part to damage to the cortex-lytic enzyme CwlJ. (v) Increases in T lag and ⌬T release were also observed when wet-heat-treated B. subtilis spores were germinated with the nonnutrient dodecylamine, while the change in I release was less significant. (vi) The effects of wet-heat treatment on nutrient germination of B. cereus and B. megaterium spores were generally similar to those on B. subtilis spores. These results indicate that (i) some proteins important in spore germination are damaged by wet-heat treatment, (ii) the cortex-lytic enzyme CwlJ is one germination protein damaged by wet heat, and (iii) the CaDPA release process itself seems likely to be the target of wet-heat damage which has the greatest effect on spore germination.

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Kinetics of Germination of Wet-Heat-Treated Individual Spores of Bacillus Species, Monitored by Raman Spectroscopy and Differential Interference Contrast Microscopy

Wang

Zhang

Setlow

et al. 2011

Appl Environ Microbiol

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“…Figure 2 shows the average Raman spectra of 20 B. cereus spores from an untreated population (Fig. 2, curve a) and a population heat treated in water at 80°C for 20 min ( are from CaDPA (19,21); the band at 1,004 cm Ϫ1 is due to phenylalanine in spore protein; and the bands at 1,655 and 1,668 cm Ϫ1 are associated with ␣-helical and irregular structures of protein amide I (peptide bond CAO stretch), respectively (20). The Raman spectra also had a weak band at 1,155 cm Ϫ1 that corresponds to the C-C stretch vibration of carotenoids that is a prominent band in B. megaterium spores (3,26,32).…”

Section: Resultsmentioning

confidence: 99%

Characterization of Wet-Heat Inactivation of Single Spores of Bacillus Species by Dual-Trap Raman Spectroscopy and Elastic Light Scattering

Zhang

Kong

Setlow

et al. 2010

Appl Environ Microbiol

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Dual-trap laser tweezers Raman spectroscopy (LTRS) and elastic light scattering (ELSBacterial spores of Bacillus species are formed in sporulation and are metabolically dormant and extremely resistant to a variety of harsh conditions, including heat, radiation, and many toxic chemicals (37). Since spores of these species are generally present in foodstuffs and cause food spoilage and food-borne disease (37, 38), there has long been interest in the mechanisms of both spore resistance and spore killing, especially for wet heat, the agent most commonly used to kill spores. The killing of dormant spores by wet heat generally requires temperatures about 40°C higher than those for the killing of growing cells of the same strain (37, 43). A number of factors influence spore wet-heat resistance, with a major factor being the spore core's water content, as spores with higher core water content are less wet-heat resistant than are spores with lower core water (15,25). The high level of pyridine-2,6-dicarboxylic acid (dipicolinic acid [DPA]) and the types of its associated divalent cations, predominantly Ca 2ϩ , that comprise ϳ25% of the dry weight of the core also contribute to spore wet-heat resistance, although how low core water content and CaDPA protect spores against wet heat is not known. The protection of spore DNA against depurination by its saturation with a group of ␣/␤-type small, acid-soluble spore proteins also contributes to spore wet-heat resistance (14,23,33,37).Despite knowledge of a number of factors important in spore wet-heat resistance, the mechanism for wet-heat killing of spores is not known. Wet heat does not kill spores by DNA damage or oxidative damage (35, 37). Instead, spore killing by this agent is associated with protein denaturation and enzyme inactivation (2, 7, 44), although specific proteins for which damage causes spore death have not been identified. Wet-heat treatment also often results in the release of the spore core's large depot of CaDPA. The mechanism for this CaDPA release is not known but is presumably associated with the rupture of the spore's inner membrane (7). In addition, the relationship between protein denaturation and CaDPA release is not clear, although recent work suggests that significant protein denaturation can occur prior to CaDPA release (7). Almost all information on spore killing by moist heat has been obtained with spore populations, and essentially nothing is known about the behavior of individual spores exposed to potentially lethal temperatures in water. Given the likely heterogeneity of spores in populations, in particular in their wetheat resistances (16,18,39,40), it could be most informative to analyze the behavior of individual spores exposed to high temperatures in water.Raman spectroscopy is widely used in biochemical studies, as this technique has high sensitivity and responds rapidly to subtle changes in molecule structure (1, 22, 31). In addition,

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“…Tryptophan and phenylalanine exhibit a characteristic band at 1007 cm -1 due to breathing modes of the benzene and pyrrole rings. 31,33,34 In summary it can be said that the main differences between female and male feather pulp UVRR spectra result from different DNA contents of both genders. The Raman difference signals (see Figure 1) being not characteristic for either female or male feather pulp and therefore not useful for a gender determination arise in the range of 1450 and 1350 cm -1 , between 1150 and 1010 cm -1 , and below 1000 cm -1 .…”

Section: Resultsmentioning

confidence: 96%

Minimal Invasive Gender Determination of Birds by Means of UV-Resonance Raman Spectroscopy

et al. 2008

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The identification of avian gender is important for prosperous breeding of birds. Since birds do not possess external genital organs, endoscopic investigations, blood analysis, and molecular biological methods are applied to determine the gender in monomorphic species. However, anesthesia and blood sampling impose stress on the examined bird and should be avoided in terms of animal protection. Here we report on the application of UVresonance Raman spectroscopy as a minimal invasive method for gender determination of birds via an evaluation of feather pulp samples. Sample preparation for this investigation method is simple and facilitates a quick and easy analysis. The UV-resonance Raman spectra of the feather pulp sample extracts are dominated by DNA and protein signals. The different DNA content in male and female chicken allows for gender differentiation via its characteristic Raman fingerprint. The classification either to male or female chicken is ideally accomplished by support vector machines due to the fact that no unknown classes are involved. Recognition rates of about 95% were compared to less effective results of the unsupervised hierarchical cluster analysis. Within the scope of our investigations, principal component analysis was also applied to determine the important spectral regions for the classification of chicken's feather pulp samples.

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Raman Spectroscopy of Proteins

Abstract: The sections in this article are Introduction General Aspects of Ordinary R aman Spectra of Proteins Amides I and III Frequencies and Secondary Structures T rp … Show more

Cited by 41 publications

References 217 publications

Kinetics of Germination of Wet-Heat-Treated Individual Spores of Bacillus Species, Monitored by Raman Spectroscopy and Differential Interference Contrast Microscopy

Kinetics of Germination of Wet-Heat-Treated Individual Spores of Bacillus Species, Monitored by Raman Spectroscopy and Differential Interference Contrast Microscopy

Characterization of Wet-Heat Inactivation of Single Spores of Bacillus Species by Dual-Trap Raman Spectroscopy and Elastic Light Scattering

Minimal Invasive Gender Determination of Birds by Means of UV-Resonance Raman Spectroscopy

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