Prof.Kurt Wüthrich(Kurt Wüthrich教授 (2002年诺贝尔化学奖获得者)

Kurt Wuthrich and NMR of biological macromolecules
Nuclear magnetic resonance (NMR) spectroscopy is the only experimental technique that can determine the structures and dynamics of biological macromolecules and their complexes in solution and with atomic resolution. The award of the 2002 Nobel Prize in Chemistry to Kurt Wuthrich of the Swiss Federal Institute of Technology and The Scripps Research Institute honors his pioneering efforts in developing and applying this technique. Wuthrich shared the prize with John B. Fenn and Koichi Tanaka, who were recognized for the development of ionization methods for the analysis of proteins using mass spectrometry.

Nobel Lecture-Prof. Wuthrich
Nobel lecture by Prof. Wuthrich, a pioneer in protein structure determination by
NMR spectroscopy. Extensions of solution NMR spectroscopic methods for study of
large molecules.

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2002年諾貝爾化學獎－－生物巨分子分析
2002年諾貝爾化學獎頒給對發展生物巨分子的鑑定與結構分析之方法有重大貢獻的John B. Fenn、田中耕一和Kurt Wuthrich。前兩者發展出生物巨分子的質譜分析，後者發展出解蛋白質結構的NMR。

43歲的田中耕一是諾貝爾化學獎創設以來最年輕得主，也是繼小柴昌俊贏得諾貝爾物理學獎後，今年第二位獲得諾貝爾獎的日本人。

瑞典皇家學院為表揚三人研發解析大分子的強大工具，讓研究如蛋白質等較大分子變得可能，協助人類進一步理解生命過程，決定頒授三人諾貝爾化學獎。

三位科學家帶來的新技術不僅在新藥研發上掀起革命，還證明可應用於癌症和瘧疾的早期診斷上。利用他們的研究結果，化學家現在能迅速確實的辨識蛋白質，製造出溶液中蛋白質分子的三度空間影像，讓他們更加清楚蛋白質在細胞中的作用。三人研發的技術還可運用於運動禁藥監督、環境汙染分析、食品控管改善，未來還可能協助醫藥界找到治癌藥物。

85歲的Fenn自耶魯大學退休後，任教於Virginia Commonwealth University；田中耕一是京都精密儀器製造商島津製作所的工程師；64歲的Wuthrich在瑞士蘇黎世Swiss Federal Institute of Technology任教，並且是加州Scripps Research Institute的訪問教授。

諾貝爾化學獎獎金約一百萬美元，Wuthrich將獲得其中的五十萬美元，剩餘的五十萬美元由田中耕一與Fenn均分。

Fenn與田中是在八○年代各自獨立地為質譜儀（Mass Spectrometry）的解析技術帶來突破。Fenn發展出的「電噴霧離子化法」（electrospray ionisation），讓水份蒸發後會留下帶電的蛋白質溶液的小液滴，最後剩下蛋白質離子，然後其質量可以因其在一定距離內到達偵測器的時間來決定。田中耕一則研發出一種「輕雷射脫著法」（soft laser desorption），將細胞與組織內的各類蛋白質，以雷射迅速正確的解析出來

Wuthrich的貢獻在於他改善核磁共振（nuclear magnetic resonance, NMR）技術，證明它也可以用來研究生物巨分子，讓化學家能在溶液中解析蛋白質的構造，等於提供科學家一個與活細胞類似的環境研究蛋白質。

Wuthrich對獲獎感到驚喜，他是第五位獲得諾貝爾化學獎的瑞士人，也是自 1991 年來的第一人。

NMR SPECTROSCOPY AND BIOMOLECULAR STRUCTURE
The development of multidimensional Nuclear Magnetic Resonance (NMR) spectroscopy in recent years resulted in exploiting this technique for the determination of three dimensional structures of proteins. In fact the Nobel prize for the year 2002 in chemistry is given to Prof. Kurt Wuthrich for his work on “Three dimensional Structure of Biomolecules”.

The biggest advantage of using NMR spectroscopy is that one can obtain the 3D structure in water, which is the solvent in which most biological reactions take place (enzymes and drugs interact in water). Moreover it gives the dynamic information of the molecule. It is not that NMR spectroscopy can give all the information. It has its own limitations. As the protein size becomes bigger and bigger, the resonance assignments are not very easy, although part of the problems are overcome in recent years by tremendous development in experimental schemes. Recent published work is on a protein of 81.4 kD having nearly 723 residues (J. Am. Chem. Soc. 124, 10023-35 (2002).

Introduction to NMR Spectroscopy
Structures of biomolecules
How we can use NMR to determine these structures? What is the procedure?
COSY : Abbreviation for COrrelated SpectroscopY
TOCSY : Abbreviation for TOtal Correlated SpectroscopY
NOESY : Abbreviation for Nuclear Overhauser Effect SpectroscopY

NOESY is one of the most useful experiment as it is used to correlate the nuclei through space. This gives qualitative and quantitative information on the proximity of two nuclei. Also by measuring the intensity of the peaks, the distance information can be obtained. In this experiment after magnetization evolves during t1 period, exchange of the magnetization takes place during the mixing time through a dipole-dipole relaxation mediated relaxation.

Structure determination
All structural information is available once firm assignments of the residues are made using 2D technique. This is the crucial step. It is necessary not only to identify every amino acid type but also by its position in the polypeptide sequence. The chemical shift positions of Ca proton chemical shifts are very useful and secondary structure has characteristic pattern in this region. a helices have little chemical shift dispersion whereas b sheet have more dispersion. Second important step is to NOE parameters. It is most useful for tertiary structure determination.

Typical flow chart followed for tertiary structure determination

Sequence specific assignments
Make use of experiments like COSY, TOCSY, DQFC, etc..

Collect restraints from NOE
Make use of experiments like NOESY, ROESY, etc…

Generally other constraints are also used like Hydrogen bond, Torsion constraints, etc. (I am not going into details of all these due to limited time)

Secondary structure determination
Generally characteristic patterns exist for secondary structure in finger pring region

The constraints obtained are used in certain distance geometry algorithm to obtain tertiary structure
Refinement of the structure using molecular dynamics / molecular modelling
The procedure for the determination of structure of biomolecule is same whether it is polypeptide or protein. So far what I said is the preliminary steps. In obtaining the real structure, one has to carry out several more experiments designed to retrieve information. I am not covering all of them due to lack of time. Depending on the system one can choose several other sophisticated experiments involving multi dimensions in structure determination.

以上是节选,全文http://www.fbae.org/Channels/tools_in_biotechnology/nmr.htm

NMR专题讲座
Special Topics in NMR Spectroscopy
NMR. Software development continues to improve data acquisition, processing.
and analysis. Kurt Wuthrich shared the 2002 Nobel Prize in Chemistry for his.
NMR-based contributions to structural biology. The growth of NMR continues.
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