Plasma protein turnover and tissue exchange; influence of dietary protein and protein depletion

The rate of plasma protein turnover is more rapid in dogs receiving adequate dietary protein than when a diet devoid of protein is fed. Both albumin and combined globulins are involved in this change. The difference in turnover is reflected in a total protein half-life of 4.8 days with protein feeding versus 7.8 days without protein in the diet and in the metabolism of 1.0 and 0.65 gm. per kilogram of body weight per day on the respective diets. Additions of dietary protein from 10 to 30 per cent caused no further increase in the rate of plasma protein turnover. With protein depletion due to plasmapheresis and a very low protein diet there is evidence of reduced protein metabolism as indicated by nitrogen retention as well as a reduction in total plasma protein breakdown and interchange of isotope between plasma and tissue proteins. Following introduction of labeled plasma protein into the circulation the net amount of isotope transferred to tissues has been computed from the difference between total plasma protein breakdown and combined C¹⁴ excretion in urine and expired air. In animals receiving adequate dietary protein, tissue transfer amounts to 70 per cent of the total lost from the plasma proteins each day while the percentage rises to 85 in depleted dogs deprived of protein. In dogs with both plasma and tissue proteins labeled it can be estimated that, under conditions of protein feeding, an amount of C¹⁴ approximately equal to that lost from the plasma must recycle to account for the observed decrease in Apparent plasma protein turnover rate, (t½ of 15 versus 5 days). Without protein in the diet the isotope contribution of the tissues to the maintenance of plasma protein levels must be as great as or greater than that transferred in the opposite direction.     

腺苷酸活化蛋白激酶与骨骼肌蛋白质降解

背景:机体运动时骨骼肌收缩,ATP被大量消耗,产生大量腺苷一磷酸,导致腺苷酸活化蛋白激酶的激活。目的:综述不同运动过程中腺苷酸活化蛋白激酶活性的变化,以及腺苷酸活化蛋白激酶对骨骼肌蛋白质降解的研究成果。方法:检索中国期刊网、维普期刊数据库、www.ncbi.nlm.nih.gov/pubmed和http://highwire.stanford.edu/网站与腺苷酸活化蛋白激酶、运动、蛋白质降解研究相关的文章。并对腺苷酸活化蛋白激酶的结构与作用,不同运动过程中腺苷酸活化蛋白激酶活性的变化,以及腺苷酸活化蛋白激酶升高对骨骼肌蛋白质降解的内容进行分析综述。结果与结论:共纳入相关文献35篇。本文综述了腺苷酸活化蛋白激酶的结构、作用的研究进展;在抗阻运动和中到大强度的周期运动中,腺苷酸活化蛋白激酶活性都可能升高,而在小强度周期运动过程中腺苷酸活化蛋白激酶活性可能不升高;腺苷酸活化蛋白激酶的活化可能对骨骼肌蛋白质的降解有促进作用。     

腺苷酸活化蛋白激酶与骨骼肌蛋白质降解

背景：机体运动时骨骼肌收缩,ATP被大量消耗,产生大量腺苷一磷酸,导致腺苷酸活化蛋白激酶的激活。目的：综述不同运动过程中腺苷酸活化蛋白激酶活性的变化,以及腺苷酸活化蛋白激酶对骨骼肌蛋白质降解的研究成果。方法：检索中国期刊网、维普期刊数据库、www.ncbi.nlm.nih.gov/pubmed和http：//highwire.stanford.edu/网站与腺苷酸活化蛋白激酶、运动、蛋白质降解研究相关的文章。并对腺苷酸活化蛋白激酶的结构与作用,不同运动过程中腺苷酸活化蛋白激酶活性的变化,以及腺苷酸活化蛋白激酶升高对骨骼肌蛋白质降解的内容进行分析综述。结果与结论：共纳入相关文献35篇。本文综述了腺苷酸活化蛋白激酶的结构、作用的研究进展;在抗阻运动和中到大强度的周期运动中,腺苷酸活化蛋白激酶活性都可能升高,而在小强度周期运动过程中腺苷酸活化蛋白激酶活性可能不升高;腺苷酸活化蛋白激酶的活化可能对骨骼肌蛋白质的降解有促进作用。     

Specificity of protein C and protein S assays

Summary Specific tests for the measurement of protein C antigen and activity and protein S antigen are used in the clinical laboratory for the routine diagnosis of hereditary protein C and protein S deficiency. The performance of these tests is reviewed and discussed. Special attention is paid to the application of these tests for the analysis of patients on oral anticoagulant therapy. Presented at the ‘2nd International Symposium on Standardization and Quality Control of Coagulation Tests: Implications for the Clinical Laboratory’, Rome, September 28–29, 1989.     

Acquisition of inhibitor-sensitive protein kinases through protein design.

Protein phosphorylation is the major post-translational modification used by eukaryotic cells to control cellular signaling. Protein kinases have emerged as attractive drug targets because heightened protein kinase activity has been associated with several proliferative diseases, most notably cancer and restenosis. Until now, it has been very difficult to confirm the utility of protein kinases as inhibitor targets because very few small molecules that selectively inhibit one particular kinase are known. Discovery of highly specific kinase inhibitors has been slow because the protein family contains approximately 2000 members, all of which share a conserved active site fold. Recent work in several laboratories has sought to circumvent the problem of kinase structural degeneracy by engineering drug sensitivity into Src family tyrosine kinases and mitogen-activated protein kinases through site-directed mutagenesis. By introducing a unique non-naturally occurring amino acid into a conserved region of the enzyme's binding site, a target protein kinase can be rapidly sensitized to a small molecule. Introduction of the engineered kinase into a cell line or animal model should greatly expedite the investigation of protein kinase inhibition as a viable drug treatment. The purpose of this review is to summarize these recent advances in protein kinase drug sensitization.     

Activated protein C

Summary.  Protein C is a vitamin K-dependent plasma protein zymogen whose genetic mild or severe deficiencies are linked with risk for venous thrombosis or neonatal purpura fulminans, respectively. Studies over past decades showed that activated protein C (APC) inactivates factors (F) Va and VIIIa to down-regulate thrombin generation. More recent basic and preclinical research on APC has characterized the direct cytoprotective effects of APC that involve gene expression profile alterations, anti-inflammatory and anti-apoptotic activities and endothelial barrier stabilization. These actions generally require endothelial cell protein C receptor (EPCR) and protease activated receptor-1. Because of these direct cytoprotective actions, APC reduces mortality in murine endotoxemia and severe sepsis models and provides neuroprotective benefits in murine ischemic stroke models. Furthermore, APC reduces mortality in patients with severe sepsis (PROWESS clinical trial). Although much remains to be clarified about mechanisms for APC's direct effects on various cell types, it is clear that APC's molecular features that determine its antithrombotic action are partially distinct from those providing cytoprotective actions because we have engineered recombinant APC variants with selective reduction or retention of either anticoagulant or cytoprotective activities. Such APC variants can provide relatively enhanced levels of either cytoprotective or anticoagulant activities for various therapeutic applications. We speculate that APC variants with reduced anticoagulant action but normal cytoprotective actions hold the promise of reducing bleeding risk because of attenuated anticoagulant activity while reducing mortality based on direct cytoprotective effects on cells.     

Human erythrocyte protein 4.1 is a phosphatidylserine binding protein.

The aminophospholipids phosphatidylethanolamine (PE) and phosphatidylserine (PS) are the major phospholipids contained in the cytoplasmic leaflet of the human erythrocyte (RBC) plasma membrane and are largely confined to that leaflet over the entire RBC lifespan. In particular, PS, which comprises approximately 13% of total RBC membrane phospholipids, is normally restricted entirely to the cytoplasmic leaflet. However, molecular mechanisms that regulate this asymmetric distribution of phospholipids are largely unknown. We examined elliptocytic RBCs that completely lacked protein 4.1 (HE [4.1 degrees]), but contained normal amounts of all other peripheral membrane proteins, and found approximately 10% of total membrane PS was accessible in the exoplasmic leaflet of these membranes. Inside out vesicles (IOVs) derived from HE [4.1 degrees] RBCs bound fewer PS liposomes than did IOVs derived from normal RBCs. Normal IOVs that were depleted of proteins 2.1 (ankyrin), 4.1, and 4.2 bound fewer PS liposomes similar to HE [4.1 degrees] IOVs, and repletion with protein 4.1 restored PS liposome binding to control levels. Addition of purified protein 4.1 to PS liposomes resulted in saturable binding with the extent of binding being proportional to the liposome PS content. Our data suggests that human RBC protein 4.1 is a PS binding protein and may be involved in the molecular mechanisms that stabilize PS in the cytoplasmic leaflet of the human RBC plasma membrane.     

Imaging protein trafficking

The need to understand complex intracellular trafficking mechanisms from both a basic and disease-oriented perspective has stimulated considerable interest in the development of real-time microscopy tools. Recent advances in instrumentation and the development of molecular bioprobes, such as green fluorescent protein and its derivatives, have opened up a new era in our ability to perform close to real-time imaging of cellular events with high spatial and temporal resolution, and with high sensitivity. This review briefly introduces and discusses some of the systems and methodologies that are available from several manufacturers, including laser scanning and spinning disk confocal microscopy, and total internal reflectance microscopy.     

Serum protein fingerprinting

The core technologies in the rapidly expanding field of proteomics have matured to the point where quantitative measurements of thousands of proteins can be conducted, enabling truly global measurements of protein expression. This advent has brought with it the hope of discovering novel biomarkers that promise a renaissance in clinical medicine. To meet this need, many proteomic studies have focused on the identification and subsequent comparative analysis of the thousands of proteins that populate complex biological systems such as serum and tissues. A novel application of mass spectrometry has been in proteomic pattern analysis, which has emerged as an effective method for the early diagnosis of diseases. In stark contrast to 'classical' proteomics, proteomic pattern analysis relies on the pattern of proteins observed, rather than on the discrete identification of a protein. Proteomic pattern technology allows hundreds of clinical samples to be analyzed per day and promises to be a novel, highly sensitive predictive clinical tool to improve diagnostic and prognostic medicine.     

Modulation of protein kinase signaling by protein phosphatases and inhibitors

Protein tyrosine phosphatases (PTPs) form a large family of enzymes that serve as key regulatory components in signal transduction pathways. Defective or inappropriate regulation of PTP activity leads to aberrant tyrosine phosphorylation, which contributes to the development of many human diseases. In addition to controlling the phosphorylation states of protein kinase substrates, PTPs can also directly modulate protein kinase activity. Evidence suggests that PTPs can exert both positive and negative effects on a signaling pathway. Thus, further understanding of the fundamental role of protein tyrosine phosphorylation in complex and critical signal transduction pathways requires detailed studies of both the kinases and the phosphatases. In this review, we first summarize our current understanding of PTP structure and function. We then discuss the molecular basis of PTP substrate specificity, focusing primarily on mitogen-activated protein (MAP) kinase phosphatase 3. We demonstrate that the MAP kinase phosphatases display exquisite substrate specificity requiring extensive protein-protein interactions for precise down-regulation of MAP kinase activity. We also highlight our recent progress in developing small molecule PTP1B inhibitors. Using a novel combinatorial approach that is designed to target both the active site and a unique peripheral site in PTP1B, we have obtained a PTP1B inhibitor with 2.4 nM affinity and orders of magnitude selectivity against a panel of PTPs. Currently, some of the compounds are being evaluated in both cell and animal models to further define the role of PTP1B in insulin signaling.