Galactose and lactose transport inKluyveromyces lactis

Transport of lactose intoKluyveromyces lactis was accomplished by a highly specific system inducible by lactose and galactose. The biosynthesis of the transport enzyme was strongly repressed by glucose. For non-induced cells, lactose penetrated by passive transport, like galactose in any type of cells. The lactose transport showed aK _m 1.2 –4 mm, was temperature-dependent (76 kJ/mol) and was blocked by metabolic inhibitors.     

Uptake of galactose and lactose by Kluyveromyces lactis: biochemical characteristics and attempted genetical analysis

Study of the lactose and galactose transport systems in Kluyveromyces lactis has shown that lactose uptake is by active transport. The transport system is under monogenic control and is inducible. Galactose uptake is also by active transport but the system is controlled by two genes which, in the four strains we studied, are present only in K. lactis CBS 2359. Galactose uptake in the other K. lactis strains is by a simple diffusion process.     

Amylase activity inPichia polymorpha

The amylase activity ofPichia polymorpha was studied during growth. The localization of activity in the cell and the general properties of the enzyme are described. Two types of activity were observed but they could not be ascribed to two distinct enzymes.     

Glucose metabolism in the yeast Schwanniomyces castellii: role of phosphorylation site I and an alternative respiratory pathway.

Glucose metabolism in a Crabtree-negative yeast, Schwanniomyces castellii, and a cytochrome b-deficient mutant of this strain was investigated in chemostat culture. The wild-type and mutant strains exhibited the same behavior. Oxidative metabolism was observed when the substrate uptake rate (qS) was low. Fermentative metabolites were excreted when the qS value was higher than 0.40 g.g-1.h-1, indicating the occurrence of a respirofermentative metabolism; however, the respiratory quotient (RQ) remained near 1. When fermentation occurred, the cytochrome pathway was repressed but not the salicylhydroxamic acid (SHAM)-sensitive pathway. The presence of an alternative SHAM-sensitive respiratory pathway and the presence of phosphorylation site I in all metabolic conditions explained the RQ value of 1 and accounted for high biomass yields in oxidative metabolism conditions (0.62 g.g-1 for the wild-type strain and 0.31 g.g-1 for the cytochrome b-deficient mutant strain).     

Influence of culture conditions on the cell yield and amylases biosynthesis in continuous culture by Schwanniomyces castellii

Schwanniomyces castellii excreted -amylase and amyloglucosidase into the medium in the presence of starch. The biosynthesis and the rate of excretion were influenced by dissolved oxygen (specially for -amylase), pH of the culture and dilution rate. The cell yield observed (0.59) remained constant up to D=0.35h^-1 with starch as substrate. But in the case of growth on glucose, the yield observed was equal to 0.62 up to a dilution rate of D=0.18 h^-1. Beyond this value Y x/s decreased and ethanol was produced. The onset of fermentation dependend partly on the nature of the substrate and not only on the environment in particular on the quantity of dissolved oxygen present.     

Guidelines for evaluating performance of oyster habitat restoration

Restoration of degraded ecosystems is an important societal goal, yet inadequate monitoring and the absence of clear performance metrics are common criticisms of many habitat restoration projects. Funding limitations can prevent adequate monitoring, but we suggest that the lack of accepted metrics to address the diversity of restoration objectives also presents a serious challenge to the monitoring of restoration projects. A working group with experience in designing and monitoring oyster reef projects was used to develop standardized monitoring metrics, units, and performance criteria that would allow for comparison among restoration sites and projects of various construction types. A set of four universal metrics (reef areal dimensions, reef height, oyster density, and oyster size–frequency distribution) and a set of three universal environmental variables (water temperature, salinity, and dissolved oxygen) are recommended to be monitored for all oyster habitat restoration projects regardless of their goal(s). In addition, restoration goal‐based metrics specific to four commonly cited ecosystem service‐based restoration goals are recommended, along with an optional set of seven supplemental ancillary metrics that could provide information useful to the interpretation of prerestoration and postrestoration monitoring data. Widespread adoption of a common set of metrics with standardized techniques and units to assess well‐defined goals not only allows practitioners to gauge the performance of their own projects but also allows for comparison among projects, which is both essential to the advancement of the field of oyster restoration and can provide new knowledge about the structure and ecological function of oyster reef ecosystems.     

Proline-Rich Salivary Proteins Have Extended Conformations

Three basic proline-rich salivary proteins have been produced through the recombinant route. IB5 is a small basic proline-rich protein that is involved in the binding of plant tannins in the oral cavity. II-1 is a larger protein with a closely related backbone; it is glycosylated, and it is also able to bind plant tannins. II-1ng has the same polypeptidic backbone as II-1, but it is not glycosylated. Small angle x-ray scattering experiments on dilute solutions of these proteins confirm that they are intrinsically disordered. IB5 and II-1ng can be described through a chain model including a persistence length and cross section. The measured radii of gyration (R_g = 27.9 and 41.0 ± 1 Å respectively) and largest distances (r_max = 110 and 155 ± 10 Å respectively) show that their average conformations are rather extended. The length of the statistical segment (twice the persistence length) is b = 30 Å, which is larger than the usual value (18 Å − 20 Å) for unstructured polypeptide chains. These characteristics are presumably related to the presence of polyproline helices within the polypeptidic backbones. For both proteins, the radius of gyration of the chain cross-section is R_c = 2.7 ± 0.2Å. The glycosylated protein II-1 has similar conformations but the presence of large polyoside sidegroups yields the structure of a branched macromolecule with the same hydrophobic backbone and hydrophilic branches. It is proposed that the unusually extended conformations of these proteins in solution facilitate the capture of plant tannins in the oral cavity.     

Overexpression and characterization of two human salivary proline rich proteins

Proline rich proteins (PRP) are among major human saliva constituents and are known to interact with wine tannins that are involved in astringency. To characterize these interactions, a human salivary proline rich pro-protein, PRB4S, was overexpressed in Pichia pastoris. Six recombinant proteins resulting from maturation in bioreactor were detected by SDS-PAGE analysis between 15 and 45 kDa (apparent molecular weight). Two of them, the 45 and the 15 kDa ones, were isolated from culture supernatant by adsorption and permeation chromatography. They were characterized by N-terminal sequencing and MALDI-TOF analysis after trypsic digestion. The 45 kDa protein is glycosylated while the 15 kDa one was obtained after a furin-like proteolysis. Both of them are similar to human whole saliva PRP resulting from proteolysis of PRB4S pro-protein in Golgi network and known as II-1 and IB-5. Because of their sensitivity to proteolysis or their unusual mobility on SDS-PAGE gel, these recombinant proteins seem to be intrinsically unstructured proteins.     

Influence of culture conditions on the production of heterologous interleukin 1β by Kluyveromyces lactis

Summary Under the control of the repressible PHO5 promoter, the expression of gene encoding interleukin 1 (Il1) was derepressed when the medium was depleted of free inorganic phosphate (Pi). Maximum heterologous protein synthesis was obtained in the presence of 75 mg KH₂PO₄/1 (for 20 g glucose/l). The successful heterologous protein production greatly depends on nutritional culture conditions as Il1 production efficiency was increased by 83% through optimization of the growth medium. Comparison of different phosphate-limited cultivation strategies led to the development of a batch culture procedure with nutrient pulses to delay induced oxido-fermentative glucose metabolism and increase the Il1 production to 135 mg/l.