v********a
发帖数: 646 | 1 我需要分析多种组织和细胞里面的RNA-protein interactions, 但是市面上有多种
Protease Inhibitor Cocktail。请问哪种cocktail，适合于RBP的免疫沉淀？
EMD Millipore protease inhibitor cocktail
SIGMAFAST protease inhibitor tablets
BD PharMingen protease inhibitor cocktail
Roche Complete protease inhibitor cocktail
Nacalai Tesque protease inhibitor mixture
Thermo Scientific Halt Protease Inhibitor Cocktail
http://www.biocompare.com/pfu/120065/soids/2254598-2254612/Inhi |
|
k******0
发帖数: 1073 | 2 你的buffer有没有protease inhibitors，如PMSF， pepstatin等，它们是Tev
protease的抑制剂。有没有DTT，通常我们做Ni－NT不加DTT，好像TEV protease最好有
些DTT存在。还有，in column proteolysis
通常不如in solution digestion有效，多加些protease。 TEV protease特异性很强，
加多了也不怕。如果可以，还可以在20C水解，比4C有效。 |
|
S*******l
发帖数: 4637 | 3 3C protease inhibitor是抑制鼻病毒蛋白酶的。SARS的蛋白酶和3C protease 很像，
3C protease inhibior对SARS 病毒也有效。
上次没赶上试用，这次应该给重症病人使用。 |
|
m*********r
发帖数: 49 | 4 正在纯化一个his-tag的蛋白。我想让蛋白结合到Ni柱上后用TEV protease剪切后收集
切下来的蛋白，我觉得这样比用immidazole洗脱纯度高，还省了以后再切tag的麻烦。
现在问题是，老板买了个qiagen的fast start kit (gravity flow的那种）， 里面的
Ni-agarose成型了之后像gel一样，不能被重新悬浮，我在加TEV protease洗脱的时候
（4C 过夜），溶液不能完全浸没Ni-agarose，跑胶后和immidazole洗脱的比较发现洗
脱效率很低，我怀疑是因为TEV protease不能很好地与Ni-agarose接触造成的。
请教下有什么推荐的解决方法么？别家卖的Ni-NT好像有那种纯化过程中一直都自由悬
浮的，
不知道哪家的好，或者我的TEV proteasejianqie剪切加洗脱的方案有设么可以改进的
地方？多谢！ |
|
k****x
发帖数: 2751 | 5 需要protease 和 phosphatase都可以买到。 |
|
m******t
发帖数: 109 | 6 sigma protocol does not mention protease inhibitor at all (i use sigma M2
beads).
and samples need to rotate at RT for 1 hour.
that is why i asked |
|
m*********r
发帖数: 49 | 7 1.Protease inhibitor我只有在裂解细胞的时候加入的，不过在洗脱之前已经wash了几
次，洗脱的时候应该没什么残留了。
2. 我没加DTT,刚看了下别的protocol，貌似也推荐用1 mM DTT，我下次加入试试。（-
_- 第一次试没经验）
3. TEV加入量的问题也值得我多算计一下，多谢回复！ |
|
m*********D
发帖数: 1727 | 8 我们用Roche Complete protease inhibitor cocktail 小丸子，方便。 |
|
w*****n
发帖数: 107 | 9 any EDTA-free protease inhibitor is OK |
|
k*****n
发帖数: 323 | 10 我们用的是rick young 的protocol。
Lysis Buffer 1 Add protease inhibitors just before use, filter and keep cold
. Consists of 50 mM HEPES-KOH, pH 7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol,
0.5% NP-40, 0.25% Triton X-100, 1× protease inhibitors Lysis Buffer 2 Add
protease inhibitors just before use, filter and keep cold. Consists of 10 mM
Tris-HCl, pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1× protease
inhibitors
Lysis Buffer 3 Add protease inhibitors just before use, filter and keep cold
. Consists of... 阅读全帖 |
|
z*t
发帖数: 863 | 11 Beutler得奖，Medzhitov会不会有意见？
http://www.sciencedirect.com/science/article/pii/S1074761309002
Ruslan Medzhitov
This year marks the 20th anniversary of a publication (Janeway, 1989) that
in many ways revolutionized our understanding of the immune system. As part
of the proceedings of the Cold Spring Harbor Symposium on Immune Recognition
, the paper authored by the late Charles A. Janeway, Jr. was not a standard
peer-reviewed publication. In fact, Charlie used to refer to it as “the
best paper I've ... 阅读全帖 |
|
S*******l
发帖数: 4637 | 12 这个SARS-CoV protease inhibitor很多文章了，很多实验室都搞出来过。
蛋白酶抑制剂很成熟的方法了。他主要解了SARS-CoV protease的结构，就是一公颜宁
搞的那种。这样，活性中心和底物结构就很清楚了，可以看着结构设计抑制剂，就是
structure based drug design.
结构化学的价值体现之一。
可以合成出非常有效的抑制剂。
只不过野生型SARS被土共从地球上消灭了，这种高效抑制剂就没有用武之地，没有商业
价值了，所以没有制药公司跟进搞成药上市。
假如SARS-Cov protease inhibitor已经成药，会是治疗的特效药，阻断病毒组装，切
断病毒复制，大大减少viral load, 给免疫系统胜利的机会。还可以制成气雾剂用于预
防。
可惜，现在不赶趟，他最对带些实验室制备的少量对有限的病人治疗。
目前，最现实的，是使用特异性差一些的HIV protease inhibitor, 对SARS-CoV也有抑
制作用。
国内医院已经在用了，ritonavir, lopinavir |
|
S*******l
发帖数: 4637 | 13 这样也行？那我也可以邀功了，第一个提出抗艾滋病protease inhibitor防治新冠。
http://www.mitbbs.com/article_t/Military/55627213.html
3C protease inhibitor是抑制鼻病毒蛋白酶的。SARS的蛋白酶和3C protease 很像，
3C protease inhibior对SARS 病毒也有效。
上次没赶上试用，这次应该给重症病人使用。
nelfinavir是上市的抗艾滋病药物，对SARS复制有抑制作用，可以直接上了。
加到用药方案里。 |
|
s*******2
发帖数: 598 | 14 See below 1997 paper
Biochem J. 1997 Apr 1;323 ( Pt 1):233-7.
Actin is cleaved during constitutive apoptosis.
Brown SB, Bailey K, Savill J.
SourceDivision of Renal and Inflammatory Disease, Department of Medicine,
University Hospital, Nottingham NG7 2UH, UK.
Abstract
Proteases play an important role in the programme of cell death by apoptosis
but little is known of the substrates cleaved, particularly in constitutive
models of this type of cell death. Neutrophils spontaneously undergo
apoptosis ... 阅读全帖 |
|
B******1
发帖数: 9094 | 15 A widely-accepted idea is that work and play cannot possibly be used in the
same sentence. Indeed, whoever thought they were must have heard wrong.
Work is something you do so that you have enough money for a decent method
of recreation. However, that is not always the case, as studies have shown
that interest in one’s subject quickens the rate of learning, which is true
if you think about it. If you find something intriguing, you would, at
any rate, be keener to learn about it. If you believe i... 阅读全帖 |
|
l******k
发帖数: 27533 | 16 这是重大突破的药，跟以前用的protease inhibitor完全不同
是的，protease inhibitor是艾滋病药类之一
这类药副作用很大，drug drug interaction非常多
有了这个药，protease inhibitor已经不推荐用于丙肝了 |
|
p*3
发帖数: 45 | 17 Caspase-3
Dixit VM.
Yama/CPP32 beta, a mammalian homolog of CED-3, is a CrmA-inhibitable
protease that cleaves the death substrate poly(ADP-ribose) polymerase. Cell.
1995 Jun 2;81(5):801-9.
Miller, D. K.
Identification and inhibition of the ICE/CED-3 protease necessary for
mammalian apoptosis. Nature 376: 37-43, 1995. [PubMed: 7596430]
Caspase-8
Dixit VM.
FLICE, a novel FADD-homologous ICE/CED-3-like protease, is recruited to the
CD95 (Fas/APO-1) death--inducing signaling complex. Cell. 1996 Jun |
|
K******S
发帖数: 10109 | 18 Urea肯定没问题，放心。
PROTEASE INHIBITOR是个GOOD POINT。现在好多LAB都不用大分子的PROTEASE
INHIBITOR（SOY BEAN提取出来的那几种），只用小分子的。因为大分子的PROTEASE
INHIBITOR确是会CARRY OVER到最后的DIGESTION，这已经是共识了。
remove |
|
g*****p
发帖数: 451 | 19 这个问题不难回答
从你以前的帖子已经知道你找到了酶被水解的位点
那你只需要合成一段相关的底物就可以了
然后还可以标记一下荧光，加点酶，看能不能切
要是能切的话，连kcat/km都可以算出来
如果能切，一般来说trans-leavage和cis-cleavage的认定才是
最头疼的事
一般来说得需要做个核磁或者晶体结构才行
要是你能拿到zymogen的结构呢，这个是比较好回答了
但你也说了这个酶不稳定，你想拿到zymogen的结构不做几个点突变把它稳定住
是不行的
不过现在为止你的实验数据有点让我困惑的地方（我不是说你的实验数据不对，只是说很
难解释这个现象，所以希望你先再确证一下）
1.你说4度放过夜，该酶都会变成truncated，这个和你开始express出来recombinant的
protease有两者混合的现象结合起来看，很难让我相信这个酶会有autocleavage的活性
因为要知道，ecoli表达重组蛋白，其细胞中的protease浓度是很高的。如果真的具有
自剪切活性，你根本不可能拿到full-length的protease,expression的时候就自己切光
了。因... 阅读全帖 |
|
R*Q
发帖数: 179 | 20 You know the trick if you compare his father's publications to his.
Chou KC's
Graphic analysis of codon usage strategy in 1490 human proteins.
Zhang CT, Chou KC.
J Protein Chem. 1993 Jun;12(3):329-35.
PMID: 8397791 [PubMed - indexed for MEDLINE]
A vector projection approach to predicting HIV protease cleavage sites in
proteins.
Chou KC, Zhang CT, Kézdy FJ.
Proteins. 1993 Jun;16(2):195-204.
PMID: 8332607 [PubMed - indexed for MEDLINE]
A new approach to predicting protein folding types.
Chou KC, Z... 阅读全帖 |
|
n*****u
发帖数: 9 | 21 Actually the Science structure reports a protease from a related virus, but
not SARS virus. So in this sense, Rao's group does determined the first SARS
virus protease structure. But if you check PDB, the first released coordinates
of this protease structure is not by Rao's group, but by another group. I
think they determined the structures almost simultaneously, however, the other
group released first.
However, I don't like the way this news presents: "已经表达出２３４
个与人类健康密切相关的重要蛋白质，并解析出５０余个重要蛋白质的精细 |
|
S*******l
发帖数: 4637 | 22 血管紧张素抑制剂，上世纪八十年代的。众多艾滋病 protease inhibitors, 治疗
mutiple myeloma 的 proteasome inhibitor, 治疗冠状病毒的protease inhibitor。
都是明确的target 蛋白的晶体结构后针对性设计的。
你不懂就不要来现眼胡喷。 |
|
S*******l
发帖数: 4637 | 23 不懂就表露怯。
结构指导的药物设计成功例子不要太多。现在基本所有的抗艾滋的protease inhibitor
都是这么来的。
当年的SARS，要不是被中国靠防疫隔离灭绝了，也要上冠状病毒protease inhibitor特
效药了，也是解了结构才能设计出来的特效药。
还有治疗multiple myeloma 的 proteasome inhibitor,没结构能做出来么？
还有现在很多抗癌的kinase inhibitor，很多都是有结构才有的针对性设计。
你有个靶点蛋白的结构，药厂不要抢疯掉。
施和颜的成就你一辈子也无法企及，胡喷个屁。 |
|
o*******a
发帖数: 242 | 24 抗艾滋病的药克力芝是protease inhibitor, 冠状病毒2.0是RNA病毒, 复制时需要
protease. 克力芝抑制了冠状病毒的复制.
感谢王广发 |
|
v******s
发帖数: 6949 | 25 http://en.wikipedia.org/wiki/Antinutrient
Phytic acid has a strong binding affinity to minerals such as calcium,
magnesium, iron, copper, and zinc. This results in precipitation, making the
minerals unavailable for absorption in the intestines.Phytic acids are
common in the hulls of nuts, seeds and grains.
Protease inhibitors are substances that inhibit the actions of trypsin,
pepsin and other proteases in the gut, preventing the digestion and
subsequent absorption of protein. For example, Bowma... 阅读全帖 |
|
r***u
发帖数: 1272 | 26 2．方法论的转变
要想在科学研究上取得突破和成功，只有时间的付出和刻苦，是不够的。批判性分析（
critical analysis）是必须具备的一种素质。
研究生与本科生最大的区别是：本科生以吸取学习人类积累的知识为主、兼顾科学研究
和技能训练；而博士生的本质是通过科学研究来发掘创造新知识，当前和以往学习的知
识都是为了更好地服务于科学研究。在以学习知识为主的本科生阶段，提出问题固然重
要，但答案往往已经存在，所以问题是否critical没有那么关键。博士生阶段则完全不
同，必须具备critical analysis的能力，否则不可能成为优秀的科学家。这一点，我
称之为方法论的转变。
其实，整个大学和研究生阶段教育的实质就是培养critical analysis的能力，养成能
够进行创新科研的方法论。这里的例子非常多，覆盖的范围也非常广，在此举几个让我
终生难忘的例子。
(1) 正确分析负面结果（negative results）是成功的关键。
作为生命学科的一名博士生，如果每一个实验都很顺利、能得到预料中的正面结果（
positive results），除个别研究领域外，一般只需要... 阅读全帖 |
|
s******y
发帖数: 28562 | 27 This is a convenient recipe: 0.5% Triton X-100, 20~50mM Hepes-KOH, pH 7.8,
50mM KCl, 2mM MgCl2, 5mM EGTA,1mM EDTA, 100mM Sucrose, Plus cocktail of
protease inhibitors.
Triton X-100 is for lysis of cell membrane, Sucrose is for the stablity of
most proteins, protease inhibitors is again proteolysis.
However, this may differe depend on your purpose. You should check related
reference to get a recipe that works for your experiment
By the way, in some recipe, I notice they don't use Triton, don't un |
|
l*******k
发帖数: 361 | 28 先透析，去掉imidazole， glycerol 5-10%, protease inhibitor不是必须的，可以加
一点EDTA, 1mM, 绝大部分serine protease没有Mg离子都没有活性了，有的蛋白要加DTT或者beta mecapitalmethanol,保持溶液的还原状态，视情况而定。
然后分装，用液氮进行迅速冷却，然后保存在-80, 有些比较脆弱的蛋白即使这样保存也会失活，那就需要找到合适的buffer，或者每次都用fresh made的蛋白
推荐你去读一下一个科大师兄写的从纯化到星辰，里面提到了很多蛋白提纯跟保存的技
术，对于做蛋白的很有帮助 |
|
s******y
发帖数: 28562 | 29 NO. Trypsin is a protease and won't bother with RNA.
Also, in most RNA extraction kit, there will be a step to denature all the
protein, that includes protease. |
|
e****s
发帖数: 1125 | 30 如果我是你，先检查下面2个问题再考虑Protease的问题。Protease也不会切得干干净
净，你应该在胶上看到略小的带才是。
1.tag有没有切下来？
Ni-column用咪唑洗一下，跑跑看柱子上挂的是不是只有小tag.
2.没Tag了，蛋白是不是沉淀了？
酶切前后的样品高速离心一下，上清和pellet都测一下。
70aa |
|
s********n
发帖数: 2939 | 31 他们使用polyArg和polyGlu，中间用protease的特异位点链接，当protease切下后，
Cy5标记的polyArg就和肿瘤细胞结合标记了，所以不用抗体。 |
|
s*******e
发帖数: 1010 | 32 Do u have a cleavable His-tag on you protein? Sometimes protein (esp
membrane protein) has a low yield which can not be found by comparing
induced and non-induced samples.
If your protein is really important for you, try add a Thrombin, TEV or SUMO
protease cutable His-tag and do a simple separation with Nikel beads. Then
check if there is difference between SDS-PAGEs with and without protease
incubation. Good luck.
IPTG), spin |
|
s******y
发帖数: 28562 | 33 Merck公司推出的新药Boceprevir对丙型肝炎Hepatitis C病毒有很好的抑制效果。
这个药最早是由 Schering-Plough开发的，这个公司在２００９年被 Merck 买下.
Boceprevir 属于蛋白酶抑制剂，特异性的抑制hepatitis C virus 的一个蛋白酶NS3
protease ，而导致病毒蛋白不能被完全处理而无法组装和成熟.
由于丙种肝炎病毒的寄主特异性和在体内的感染过程的特殊性。该病毒目前没有有效的
体外细胞培养系统，也缺乏可以大规模筛选的动物模型，所以无法采用小分子筛选方式
。该药的开发几乎是纯粹由蛋白结构研究和binding prediction 推导开的，在对NS3
protease进行结晶之后，研究组在一些已知信息的基础上，对蛋白酶的活性基团serine 的侧链进行了抑制物设计
。在得出了最初的蛋白多肽lead 之后进行了进一步的优化，
最终得出了一个最有效的分子结构。
Federal health officials say a highly-anticipated drug to treat hepatitis C
made ... 阅读全帖 |
|
s******y
发帖数: 28562 | 34 蛋白降解一般而言可以至少有两个途径：proteasome, lysosome, 再加上limited
proteolysis by calpain or other proteases.
前者可以用MG132 or beta-lactone
中间可以用100mM NH4Cl
后者可以用calpain inhibitors
MG132 本身是一个小多肽模拟物，一般认为是堵塞chymotrypsin-like cleavage
activity. 所以其实也不是一个很干净的inhibitor, cross-react with calpain and
other protease. 但是它在浓度比较高的时候(20uM) 对proteasome 抑制
是非常明显的。
beta-lactone 主要通过和proteasome subunit crosslinking 而起作用，比
MG132 专一性稍微好一点点，但是抑制作用不是特别明显。
不管哪个inhibitor 都不能完全解决问题。我不能说你的导师到底是对还是错，
但是你应该找其他方法来佐证。比方说，你可以看看蛋白用这个东西处理的时候，
... 阅读全帖 |
|
l******g
发帖数: 1623 | 35 怎么样证明一个protein可以self cleavage, 而不是其他的protease协助。这蛋白在体
内也是会cleaved，没人得到过full length的，文献上说是其他protease切的。我做的
recombinant, 无论真核还是大肠杆菌表达出来的，都不稳定，纯化出来的都是full
length 和 truncated 在一起。4度放放，过夜都会变成truncated. 我怀疑这蛋白自己
切了自己，不知道怎么证明。 |
|
i***0
发帖数: 160 | 36 If you dilute the high concentration protease to low concentration its
activity increase, then the decreased activity at high concentration is
reversible, which is good. It might due to the storage buffer you use for
concentration not friendly for the activity of the enzyme, or reversible
oligomerization that enclose the active site.
If you dilute the high concentration protease and can not restore the
activity you have seen while it is at low concentration, it is possible,
that it is aggregated... 阅读全帖 |
|
R*Q
发帖数: 179 | 37 http://chou.med.harvard.edu/jameschou_CV.htm
Dr. James Chou
Education
1993 B.S. in Physics, University of Michigan, Ann Arbor
1994-1999 Ph.D. in Biophysics, Harvard University, Boston
Advisor: Professor Gerhard Wagner
46.
A formulation for correlating properties of peptides and its application to
predicting human immunodeficiency virus protease-cleavable sites in proteins.
Chou JJ.
Biopolymers. 1993 Sep;33(9):1405-14.
PMID: 8400033 [PubMed - indexed for MEDLINE]
Related citations
47.
The quin... 阅读全帖 |
|
w***7
发帖数: 1637 | 38 source：http://bbs.netbig.com/thread-2491712-1-2.html
看了一下，今年不错啊，和清华并驾齐驱。
而且下半年我知道的还有两篇NS。
~~~~~~~~~~~~~~~~~
1.
Yuan W, Wu T, Fu H, Dai C, Wu H, Liu N, Li X, Xu M, Zhang Z, Niu T, Han Z,
Chai J, Zhou XJ, Gao S, Zhu B
Dense Chromatin Activates Polycomb Repressive Complex 2 to Regulate H3
Lysine 27 Methylation
Science, DOI: 10.1126/science.1225237, Aug. 24, 2012
2.
Liu B, Du H, Rutkowski R, Gartner A, Wang X
LAAT-1 Is the Lysosomal Lysine/Arginine Transporter That Maintains Amino
Acid Homeostasis
S... 阅读全帖 |
|
k******0
发帖数: 1073 | 39 It is pretty hard to predict the proteolysis of a native protein and
therefore you have to try it out yourself by using different available
proteases or even a mixture if the proteases do not affect each other. |
|
S**********e
发帖数: 1789 | 40 给你看看我们系刚进来的助理教授
Vanderbilt University School of Medicine
Fingleton, Barbara Mary , Ph.D. Assistant Professor of Cancer Biology
Publications
Barrett, CW, Fingleton, B, Williams, A, Ning, W, Fischer, MA, Washington, MK
, Chaturvedi, R, Wilson, KT, Hiebert, SW, Williams, CS. MTGR1 is required
for tumorigenesis in the murine AOM/DSS colitis-associated carcinoma model.
Cancer Res, 71(4), 1302-12, 2011.
Koon, HB, Fingleton, B, Lee, JY, Geyer, JT, Cesarman, E, Parise, RA, Egorin,
MJ, Dezube, BJ... 阅读全帖 |
|
l******a
发帖数: 3339 | 41 negative control: no transfection, no target protein detected. 证明没有內源
蛋白，
positive control: any other flag tagged protein, 证明抗体ok，
最后是同一个质粒，把gfp放进去检测，以证明你的质粒系统没问题，抗体没问题
这些都做了，估计就是protease cleavage了，我觉得不会切得那么彻底，cell lysate
加protease inhibitor了吗？ |
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S******e
发帖数: 393 | 42 将S1 (cytosolic protein) 和 S3 (nuclear soluble protein?) 去掉后，剩下的
pellet再用0.1%的Triton extract, 收集到的上清暂称为X，想知道X是些什么蛋白？
Chromatin associated protein?
方法如下：
1. Cells were extracted with Buffer A (10mM Hepes pH7.9, 10mM KCl, 1.5mM
MgCl2, 0.34mM Sucrose, 10% Glycerol, 1mM DTT plus protease inhibitors)
containing 0.1% Triton for 5 min on ice, centrifuge 1300g X5 min, discard
supernatant (S1), wash once with BufferA and discard supernatant.
2. Extract cell pellet with Buffer B (3mM EDTA, 0.2mM EGTA, 1mM... 阅读全帖 |
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d********n
发帖数: 15 | 43 it's not degradation (proteases work less efficiently at ~90C, but will be a
disaster at ~55C at which proteases have reduced activity but protein
substrates begin to open structure and give much higher accessibility), but
precipitation because proteins lose native structure under heating. This is
why SDS or LDS loading bf needs to be added before heating. 1g sds binds to
14g proteins, 10ul sds loading bf is supposed to be enough wrap all proteins
in
the sample as they are so low.
the only probl... 阅读全帖 |
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d****i
发帖数: 2346 | 44 目前在做小肠组织的western，给RIPA buffer把SDS加到2%，然后sigma的protease
inhibitor cocktail加标准量的4倍(标准量是1：100稀释用的)，但是每次lysate里面
都有蛋白降解。。。。。。。。所以想除了inhibitor cocktail还能加点啥瞬间灭火
protease还能用于western？ |
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r*****m
发帖数: 3619 | 45 和老子这一代留欧的王大成，留学期间直接贡献了一些方法学，Robert Huber 在诺贝
尔奖得奖致辞上特别提了他的贡献。当然他是一个学生。所谓晶体指导药物设计，那时
就吹起来了。
In 1970, I had begun work on the basic pancreatic trypsin inhibitor which
has later become the model compound for the development of protein NMR,
molecular dynamics, and experimental folding studies in other laboratories.
Work in the field of proteolytic enzymes and their natural inhibitors has
been continued and extended to many different inhibitor classes, proteases,
their proenzymes, and complexes betwe... 阅读全帖 |
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f*******d
发帖数: 92 | 46 不好意思，今天才看到。
你直接加SDS－loading buffer, 里面含有protease 和 phosphotase inhibitor,然后
迅速刮下来煮了。 煮完之后sonicate, spindown 取supernatant, 测浓度。 然后用含
有protease inhibitor + phosphotase inhibitor 的lysis buffer, 或者PBS啥的稀
释， 10 倍或者20倍， 保证最终SDS浓度小于0.2%。 然后加抗体 4C 1h, 然后加Beads
4C overnight. |
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发帖数: 1 | 47 No matter how the kon/koff ratio favors the complex formation, a non-
covalent binding will definitely go through a on/off equilibrium. The
Antibody-
antigen interaction may be the highest affinity (0.1-1 nM of Kd) complex
that people have ever studied in biological systems, however the equilibrium
does exist in the Ab-
Ag complex. That's how SPR analysis is working to assess the affinity.
I do not believe that non-covalent complex will last forever when the
effective
concentration of substance ... 阅读全帖 |
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b*****h
发帖数: 783 | 48 HCV的传统治疗方法是用干扰素(INF)+Ribavirin. 属于第一代吧
目前各个公司根据HCV 病毒复制途径，开发抑制病毒第二，第三代药物。而且，这些药
物可以和第一代药浑成cocktail。增加治疗效果。
HCV 复制的靶点蛋白包括 NS3, NS4A, NS4B, NS5A 和NS5B
1) Protease inhibitors (NS3/4A protease):
VRTX的Telaprevir, MRK的 Boceprevir 以及 ACHN的ACH-1625
2) Nucleoside polymerase inhibitors (NS5B):
VRUS的 RG7128, PSI-7851 以及 PSI-938 (guanine nucleotide analog).
INHX的 INX-189(guanine nucleotide analog).
IDIX的 IDX184
3) Non-nucleoside polymerase inhibitors (NS5B):
ANDS的ANA598
IDIX的 IDX-375
从最近的收购趋势，看到大公司基本都在抢2）... 阅读全帖 |
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W********g
发帖数: 610 | 49 税收五年一直处于增长状态...cash充足，我觉得风险相对于BPAX小一些
James S. J. Manuso
Thank you, Tim. Good afternoon, and thank you for joining us for Astex
Pharmaceuticals 2012 First Quarter Conference Call. During the quarter, we
made considerable operational progress on many fronts in discovery, research
and in the clinical and regulatory development of our drugs. Our balance
sheet remains sound and our financial performance was excellent. We ended
the quarter with $126 million in cash and marketable securities and we
po... 阅读全帖 |
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c*****a
发帖数: 1238 | 50 Bt protein: Cry1Ab, one of Cry proteins.
The midgut of these insects is alkaline, which allows for the activation of
the Cry proteins once the proteins are ingested.
The Cry proteins are then “dissolved into protoxins, which are cleaved by
proteases into active toxins".
The activated proteins then bind to cells in the insect’s midgut and form
pores in the cells’ outer membranes.
This leads to the swelling and destruction of midgut cells . The digestive
systems of the insects are then paralyzed |
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